• TABLE OF CONTENTS
HIDE
 Front Cover
 Title Page
 Preface
 Table of Contents
 Biological safety
 Information for researchers
 Programs
 Medical surveillance
 Appendix






Title: Biological safety manual
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 Material Information
Title: Biological safety manual
Physical Description: Book
Language: English
Creator: Division of Environmental Health and Safety, University of Florida
Publisher: Division of Environmental Health and Safety, University of Florida
Place of Publication: Gainesville, Fla.
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Bibliographic ID: UF00091476
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.

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Table of Contents
    Front Cover
        Front Cover
    Title Page
        Title Page
    Preface
        Page i
        Page ii
        Page iii
        Page iv
        Page v
    Table of Contents
        Page vi
        Page vii
        Page viii
        Page ix
        Page x
    Biological safety
        Page 1
        Page 2
        Page 3
        Page 4
        Page 5
        Page 6
        Page 7
        Page 8
        Page 9
        Page 10
        Page 11
        Page 12
        Page 13
        Page 14
        Page 15
        Page 16
        Page 17
        Page 18
        Page 19
        Page 20
        Page 21
        Page 22
        Page 23
        Page 24
        Page 25
        Page 26
        Page 27
        Page 28
        Page 29
        Page 30
        Page 31
        Page 32
        Page 33
        Page 34
        Page 35
        Page 36
        Page 37
    Information for researchers
        Page 38
        Page 39
        Page 40
        Page 41
        Page 42
        Page 43
        Page 44
        Page 45
        Page 46
        Page 47
        Page 48
        Page 49
        Page 50
        Page 51
        Page 52
        Page 53
        Page 54
        Page 55
        Page 56
        Page 57
        Page 58
        Page 59
        Page 60
        Page 61
        Page 62
        Page 63
        Page 64
        Page 65
        Page 66
        Page 67
        Page 68
        Page 69
        Page 70
        Page 71
        Page 72
        Page 73
        Page 74
        Page 75
        Page 76
        Page 77
        Page 78
        Page 79
        Page 80
        Page 81
        Page 82
    Programs
        Page 83
        Page 84
        Page 85
        Page 86
    Medical surveillance
        Page 87
        Page 88
        Page 89
        Page 90
        Page 91
        Page 92
        Page 93
        Page 94
        Page 95
        Page 96
        Page 97
    Appendix
        Page 98
Full Text


2008


olo0


Division of
Environmental Health & Safety















Biological

Safety

Manual









University of Florida
Finance and Administration
Environmental Health and Safety



Box 112190
Gainesville, Florida 32611
(352) 392-1591 phone
(352) 392-3647 fax
Email: BSO(iehs.ufl.edu
Internet: http://www.ehs.ufl.edu/bio


November 2008









Forward


By authority delegated from the University President, the Vice President for Finance and
Administration is responsible for the safety of all University facilities. Under this
authority, policies are developed to provide a safe teaching, research, service, housing
and recreational environment.

The division of Environmental Health and Safety was established in 1974 and given the
responsibility for the management of all safety practices and the administration of the
program.

The mission of the division of Environmental Health and Safety (EH&S) is to minimize
injury to faculty, staff, students and visitors and to minimize damage to University
property. Inherent in this mission is the charge to provide a safe and healthy environment
in which the University's activities can be pursued.

All applicable federal and state safety laws, rules and regulations are adopted by the
University. In order to carry out its duties and responsibilities, EH&S will adhere to
standards or codes related to safety which have been adopted and promulgated by
nationally recognized standards-setting organizations. The interpretation of safety codes
and standards is the responsibility of the Division of Environmental Health and Safety.

In order to assure an effective Environmental Health & Safety program for the University
of Florida, it is imperative that all individuals associated with the University comply fully
with the policies and procedures set forth in the manual.






Cover photos are from Henry Wellcome Laboratory for Cell Imagery, University of East Anglia.

Dr. Michael Wormstone provided the one in the upper left comer. It's a photo of FHL124
(immortalized human lens) cells labeled with an antibody to the transcription factor SMAD4 (a
component of the TGF-beta signaling pathway) and an Alexa-Fluor-488-conjugated secondary
antibody (green). The cells actin filaments were also labeled using Texas-Red-conjugated
phalloidin

Mr. David Moss contributed the one in the lower right comer of A UE-1 (immortalized mouse
inner ear epithelial) cell labeled with an antibody to alpha-tubulin microtubuless) and stained with
an Alexa-Fluor-488-conjugated secondary antibody (green). The cells nucleus was counterstained
with DAPI (blue).


Biosafety Manual


November, 2008










Policy Statement

It is the policy of the University of Florida to provide a safe working environment. The
primary responsibility for insuring safe conduct and conditions in the laboratory resides
with the principal investigator.

The UF Biological Safety Office is committed to providing up-to-date information,
training, and monitoring to the research and biomedical community concerning the safe
conduct of biological, recombinant, and acute toxin research and the handling of
biological materials in accordance with all pertinent local, state and federal regulations,
guidelines, and laws. To that end, we provide this manual as a resource, to be used in
conjunction with the CDC and NIH guidelines, the UF Select Agent Program, the
Laboratory Safety Manual and other resource materials from Environmental Health &
Safety.


Bio Safety Manual


November, 2008









Introduction


The University of Florida Biological Safety Manual is intended for use as a guidance
document for researchers and clinicians who work with biological materials. It should be
used in conjunction with the UF Laboratory Safety Manual, which provides more general
safety information. These manuals describe policies and procedures that are required for
the safe conduct of research at the University of Florida.

Responsibilities

In the academic research/teaching setting, the principal investigator is responsible for
ensuring that all members of the laboratory are familiar with safe research practices. In
the clinical laboratory setting, the faculty member who supervises the laboratory is
responsible for safety practices.

Lab managers, supervisors, technicians and others who provide supervisory roles in
laboratories and clinical settings are responsible for overseeing the safety practices in
laboratories.

Employees who work with biological materials are responsible for reading this manual,
carrying out the safety practices outlined here, and reporting any problems, accidents,
and spills to the appropriate faculty member.

Environmental Health & Safety will provide guidance, information, review,
monitoring, and training regarding biological safety programs, when appropriate. This
includes implementing registration activities for certain research projects, acting as a
consultant for departments regarding implementation and enforcement of biological
safety programs, evaluating work practices and personal protective equipment, providing
educational materials, tracking employee training, and medical monitoring.




















Biosafetv Manual November, 2008 iii











Emergency Phone Numbers


General

University Police Department

Gainesville Fire Department

Gas leak



Spills/Accidents

Asbestos

Biological or Recombinant Materials

Chemicals (laboratory)

Mercury

Pesticides

Radioactive Materials



Select Agents


Medical Emergency

Student Health Care Center

Shands Hospital Emergency Center

Shands Occupational Health Services
(blood and body fluid exposures only)

Needle Stick Injury


392-1161

395-0050

265-0250


1-866-477-6824 (OUCH)


Bio Safety Manual


392-1111

911

911


392-3392

392-1591

846-2550

392-8400

392-1904

392-7359

392-1589

392-1591
392-6369


Campus

HSC


November, 2008










Environmental Health & Safety
Phone Numbers


Biological Safety
Business Office
Chemical Waste
Director's Office
Diving Science & Safety
Facility & Fire Safety
Fire Equipment Services
Hazardous Materials Management
Industrial Hygiene
Laboratory Safety
Occupational Safety and Health
Occupational Medicine
Pest Control Services
Program Support
Radioactive Waste
Radiation Control & Rad. Services
Radiation Control Delivery Services
Radiation Control Health Center
Risk Management
Select Agents


392-1591
392-1591
392-8400
392-1591
392-1661
392-1904
392-2365
392-8400
392-3393
392-1591
392-1591
392-1591
392-1904
392-1591
392-8400
392-7359
392-8700
392-1589
392-1591
392-1591


Bldg. 179
Bldg. 179
Surge Area
Bldg. 179
Bldg. 21
Bldg. 179
Bldg. 179
Surge Area
Bldg. 104
Bldg. 179
Bldg. 104
Bldg. 179
Bldg. 175
Bldg. 179
Surge Area
Bldg. 175
Bldg. 175
HSC D8-17
Bldg. 104
Bldg. 179


Box 112190
Box 112190
Box 112725
Box 112190
Box 118205
Box 112200
Box 112200
Box 112725
Box 112195
Box 112190
Box 112195
Box 112195
Box 112205
Box 112190
Box 112725
Box 112205
Box 100252
Box 112205
Box 112195
Box 112190


Biosafety Manual


November, 2008










Table of Contents

1 -- B biological Safety.................................................................................................... 1
Principles of Biological Safety ............................................................................. 2
Laboratory Practice and Technique .................................................................... 2
Safety Equipment (Primary Barrier).................................................... ............ 3
Facility Design (Secondary Barrier) .................................................. ............ 3
Biosafety Levels ........................................ 4
A nim al F acilities............................... .............. ...... 6
C lin ical L ab oratories................... .. .......... ..... ...... .... ...... .... .... ................ 6
Importation and Interstate Shipment of Certain Biomedical Materials................... 7
Biological Safety Levels .......................................................................................... 7
B iosafety L ev el 1 ............................................. .......................... 8
B io safety L ev el 2 ...................................... ............................... ............... 9
Biosafety Level 3 ............................................ ...................... 13
Animal Biosafety Level 1 (ABSL-1)..................................................................... 18
Animal Biosafety Level 2 (ABSL-2)..................................................................... 20
A nim al B iosafety Level 3 (A B SL-3)..................................................... ....... ....... 23
A gents L ist ................................................................................................................... 30
Biological Safety Level 1 (B SL-1) ........................................................ ... .......... 30
Biological Safety Level 2 (B SL-2) ........................................ ...................... ....... 30
BSL-2 Bacterial Agents Including Chlamydia............... ..... ............ 31
B SL -2 Fungal A gents ......... ................. ......... ...................... .............. 32
B SL -2 Parasitic A gents .............. ......................................................... 32
BSL-2 Viruses ................................................. ................ 33
Biological Safety Level 3 (B SL-3) ........................................ ...................... ....... 35
BSL-3 Bacterial Agents Including Rickettsia .......................................... 35
B SL -3 Fungal A gents ...................... .. .. ................................ .............. 35
B SL -3 Parasitic A gents .............. ......................................................... 35
B SL -3 V iruses and Prions ........................................ .................... ..... 35
Biological Safety Level 4 (BSL-4) ................................................................ 36
B SL -4 B acterial A gents........................................................ .... .. .............. 36
B SL-4 Fungal A gents ........................................................ .............. 36
B SL -4 Parasitic A gents .............. ......................................................... 36
BSL-4 Viral Agents .................. ..................................................... 36
2 -- Information for Researchers ............................................................................ 38
Project Registration ............................................................................................. 39
Bio-Agent (BA) Registration....................... ...... ............................ 39
Recombinant DNA (R-DNA) Registration...................... ..... ............. 39
A cute Toxins (A T) R registration ........................................ ................... ...... 40
R egulated B biological M materials ........................................ ..................... ..... 40
T able 3: T oxin T able.................... .......... ...... ............ ........................ ................ .. 4 1
Select A gents ........................................ 43


vi November, 2008 Bio Safety Manual









Select A gents List ............................ ...... .......... .... .. .. .. .......... .. 44
Minors in Research Laboratories or Animal Facilities........................................... 46
Biological Waste Disposal Policy...................................... ................................. 47
C a te g o rie s ...................... .......... ................................................... 4 7
1) Infectious/potentially infectious/R-DNA .............................................. 47
2) N on-infectious w aste ......................................................... ......... ..... 47
3) Mixed radioactive/biohazardous waste ............................................. .. 47
4) Mixed chemical/biohazardous waste....................................................... 48
5) A nim al carcasses and m aterials.............................................. .... ................ 48
6) H um an rem ains ........................................... .. .... .... .. .......... .. 48
Packaging ............................................................... .... ..... ........ 48
1) Biohazard bags .............. ................................................ ............. 48
2) Sharp s .................. ............................ ............ .......... ..... 4 8
3) C orrugated cardboard boxes................................... ................................... 48
Labeling ............................................ ............... 48
1) D ate ............................... .......... ............... .......... 49
2) Name/Location/Phone Number ................ ............................................ 49
3) B iohazard sign .................. ............................... ..... .. .......... 49
T ra n sp o rt .................................................................................... 4 9
T ra in in g ............... ............................................................................... ......... ..... 4 9
Biological Waste Disposal Containers................................... ........................ 50
Biohazard B oxes ......................................................................... .. 50
Sharps Boxes.................................. .............. 50
B iohazard B ags .................................................................................................... 51
Autoclave Use and Testing ................................................................................... 51
A utoclave Testing ......................................................... .. .......... 51
R ecord-K keeping .................................................................... .. .. .... ........... 52
A utoclave O operating Procedures ........................................ ......................... 52
Autoclave Operation and Safety Training ......................................................... 53
A utoclave G uidelines............................ .......... ........................ ................ .. 54
D isinfectants .......................................................................................................... 55
L iq u id s .............................................................................. 5 5
A lc o h o ls .................................................................. .................................. 5 5
Chlorine com pounds .......................................... .......... ...... ........ .. 56
Formaldehyde ........... ............................... 56
Glutaraldehyde ..................................... ............ 56
Hydrogen peroxide .............. ............................................ ............. 56
Iodine and lodophors ........... .......... .. ........................ .. ........ .............. 57
Phenol and phenolic compounds ........................................................ 57
Quaternary ammonium compounds...................................... 57
Gases ................ ......... .................... ......... 57
E thylene O xide ................ ................................... .............. 57
Vapor Phase Hydrogen Peroxide..... .................... .............. 57
Chlorine D ioxide gas .............. ..... .. ...... ........................ ...................... 57


Biosafety Manual November, 2008 vii









O z o n e .................. ...................... ...... ........................................ 5 7
Formaldehyde Gas (from heating paraformaldehyde).................................. 58
Irradiation ......... .............................................. ..... .......... 58
U ultraviolet, U V radiation ......................................... ................ .............. 58
Ionizing radiation ............. ......................... ...... ...... ...... .. ......... .. 58
Electron B eam ............................. .............. 58
M icrow aves............................................ 58
D isinfectants B ibliography ............................................... ............................ 58
Shipment of Biological Materials ...................................................................... 63
Biological materials subject to shipping & transport regulations: ....................... 64
Transporting biological material within and around UF: ................................... 64
P e rm its : ........................................................................................... 6 5
Biological Safety Cabinets .................................................................................... 67
B biological Safety C abinets (B SC s)................................................. .... .. .............. 67
The Class I BSC........................................... .............. 67
The Class II B SC ..................................................... .. ............ 67
The Class II, Type A B SC ................................................... ................. 67
The Class II, Type B1 BSC .............................. 68
T he C lass II, T ype B 2 B SC .................................................... ... ................. 68
T he C lass II, T ype B 3 B SC .................................................... ... ................. 69
T he C lass III B SC .... ............................ ..................... .... ...... . ............ 69
Horizontal Laminar Flow "Clean Bench" ................. ....................................... 69
Vertical Laminar Flow "Clean Bench"............ .............................. ............. 69
O operations w within a Class II B SC ...................................... ..................... ........... 69
L laboratory H azards................... ......... ...................................... ................ .. 69
D econtam nation ........ .. ................................... .. .. ...... .... .. .......... 71
Surface Decontamination................... ........ ........................... 71
Gas Decontam ination........................................................ 71
Engineering Requirements .............. ..... ........ ....... .............. 72
Ultraviolet Lamps ... ..... ............................................ .............. 72
B SC Placem ent .... .......................... .................... ........ .. .............. 72
HEPA Filters .................................... .......................... .... ........ 72
Certification ofB SCs.................... .................................... ......................... 72
Emergency Procedures/Telephone Numbers ...................................... ........... .. 73
G en eral In fo rm action .......................................... .. ................ ......... ................. .. 74
M medical Em ergency ......... .................................... ...... ........... 74
Accidental injection, cuts, skin exposures....................................... ..... .......... 74
Splashes to face and eyes ....... ..... .............. ......... ........................... 75
A accidental ingestion .... ............................................ ........ ...... .............. 75
A nim al bites and scratches ....................................................................................... 76
B reak in/Security B reach .................................................................. ................. 77
H handling Biological Spills.................... .................................... ........... .............. 77
Spill in the biosafety cabinet................................................... .......................... 77
Spill in the centrifuge .................. ......................................... .. 77
Spill inside the laboratory ....................................................................... 78


viii November, 2008 Bio Safety Manual









Spill outside the laboratory ......................................................... .............. 79
F ire Safety ............................................................................................................. 80
Workplace Violence: ........................................... 81
H u rric a n e : ......................................................................................... 8 1
Tornadoes and other natural disasters:........................................ .............. ........ 82
3 -- Program s.................................................................................................................... 83
Animal Contact Medical Monitoring Program..................... .......... .......... ... 84
UF Bloodborne Pathogen Program ........................................................................ 86
B B P E exposure Inform ation ..................................................... ... ................. 86
4 -- M medical Surveillance........................................................................................... 87
HIV Research Laboratory Occupational Medicine Policy ................................. 88
P re-em p loy m ent ............................................................. ................ ...... .......... 8 8
C continuing em ploym ent...................................................... .......................... 88
Post-exposure prophylaxis ............................................................. ............ 88
References ................................................ 89
Immunoprophylaxis.............................................................................................. 90
Recommendations for Prophylactic Immunization of Laboratory Personnel Working
w ith Infectious Agents .................. ................................. .... .. .......... 91
Bacterial agents .................................... .......................... .......... 91
R ickettsial agents .................. .............................. ............. ......... 92
V iral ag en ts .............................................................................. 9 2
Vaccinia Immunization Policy ........................................................................... 94
U university of F lorida............................................. .. ............... ........ ................. .. 95
Health Surveillance for Personnel Working with Infectious Agents................... 96
Blood Serum Sam pling ..................................... ....................... ......... ...... 96
H health A ssessm ents .................... ............. ....................... ............. ................. 96
Exposure to Mycobacterium Tuberculosis ............................................. .......... 97
5 -- Appendix A Forms ........................................................................................... 98
R -D N A ................................................................................. 9 8
Bio-Agent........................................................ 98
Transgenic A nim als ......... .......................... ........ .......... ........... 98
A cute T oxin ........................................ 98
Project A ddendum .... .............................. ................................ .............. 98













Biosafety Manual November, 2008 ix











List of Tables


Table 1: Summary of Recommended Biosafety Levels for Infectious Agents .............. 28
Table 2: Summary of Recommended Biosafety Levels for Activities in Which
Experimentally or Naturally Infected Vertebrate Animals Are Used................... 29
Table 3: Toxin Table. ................................... .. ............ .. .......... ........ 41
Table 4: Dilutions of Household Bleach.................. ................. ........ .............. 56
Table 5: Summary and Comparison of Liquid Disinfectants (Page 1)......................... 59
Table 6: Summary of Practical Disinfectants ................................................ ............... 61
Table 7: Reprocessing Methods for Equipment Used in the Health Care Setting.......... 62


Bio Safety Manual


November, 2008






1 -- Biological Safety











Principles of Biological Safety
The following is from Biosafety in Microbiological and Biomedical Laboratories, 1999, HHS publication No. (CDC)
93-8395, Centers for Disease Control & Prevention/National Institutes of Health


The term "containment" is used in describing safe methods for
managing infectious agents in the laboratory environment where
they are being handled or maintained. The purpose of
containment is to reduce or eliminate exposure of laboratory
workers, other persons, and the outside environment to
potentially hazardous agents.

Both good microbiological technique and the use of appropriate
safety equipment provide primary containment, the protection of
personnel and the immediate laboratory environment from
exposure to infectious agents. The use of vaccines may provide
an increased level of personal protection. Secondary
containment, the protection of the environment external to the h
to infectious materials, is provided by a combination of facility
practices. Therefore, the three elements of containment include
technique, safety equipment, and facility design. The risk asses
done with a specific agent will determine the appropriate combine,


Laboratory Practice and Technique


laboratory from exposure
design and operational
laboratory practice and
sment of the work to be
ation of these elements.


The most important element of containment is strict adherence to standard
microbiological practices and techniques. Persons working with infectious agents or
potentially infectious materials must be aware of potential hazards, and must be trained
and proficient in the practices and techniques required for handling such material safely.
The director or person in charge of the laboratory is responsible for providing or
arranging for appropriate training of personnel.

Each laboratory should develop or adopt a biosafety or operations manual that identifies
the hazard that will or may be encountered, and which specifies practices and procedures
designed to minimize or eliminate risks. Personnel should be advised of special hazards
and should be required to read and to follow the required practices and procedures. A
scientist trained and knowledgeable in appropriate laboratory techniques, safety
procedures, and hazards associated with handling infectious agents must direct laboratory
activities.

When standard laboratory practices are not sufficient to control the hazard associated
with a particular agent or laboratory procedure, additional measures may be needed. The
laboratory director is responsible for selecting additional safety practices, which must be
in keeping with the hazard associated with the agent or procedure.

Laboratory personnel, safety practices, and techniques must be supplemented by


Biosafety Manual


November, 2008


T?7 --NIH 4th Edition
Siosofety in Microbiological
and Biomedical Laboratories








CDC









appropriate facility design and engineering features, safety equipment, and management
practices.

Safety Equipment (Primary Barrier)
Safety equipment includes biological safety cabinets (BSCs), enclosed containers, and
other engineering controls designed to remove or minimize exposures to hazardous
biological materials. The biological safety cabinet (BSC) is the principal device used to
provide containment of infectious splashes or aerosols generated by many
microbiological procedures. Three types of biological safety cabinets (Class I, II, III)
used in microbiological laboratories are described and illustrated in Appendix A. Open-
fronted Class I and Class II biological safety cabinets are primary barriers which offer
significant levels of protection to laboratory personnel and to the environment when used
with good microbiological techniques. The Class II biological safety cabinet also
provides protection from external contamination of the materials (e.g., cell cultures,
microbiological stocks) being manipulated inside the cabinet. The gas-tight Class III
biological safety cabinet provides the highest attainable level of protection to personnel
and the environment.

An example of another primary barrier is the safety centrifuge cup, an enclosed container
designed to prevent aerosols from being released during centrifugation. To minimize this
hazard, containment controls such as BSCs or centrifuge cups must be used for handling
infectious agents that can be transmitted through the aerosol route of exposure.

Safety equipment also may include items for personal protection such as gloves, coats,
gowns, shoe covers, boots, respirators, face shields, safety glasses, or goggles. Personal
protective equipment is often used in combination with biological safety cabinets and
other devices that contain the agents, animals, or materials being worked with. In some
situations in which it is impractical to work in biological safety cabinets, personal
protective equipment may form the primary barrier between personnel and the infectious
materials. Examples include certain animal studies, animal necropsy, agent production
activities, and activities relating to maintenance, service, or support of the laboratory
facility.

Facility Design (Secondary Barrier)
The design of the facility is important in providing a barrier to protect persons working
inside and outside of the laboratory within the facility, and to protect persons or animals
in the community from infectious agents that may be accidentally released from the
laboratory. Laboratory management is responsible for providing facilities commensurate
with the laboratory's function and the recommended biosafety level for the agents being
manipulated.

The recommended secondary barriers) will depend on the risk of transmission of
specific agents. For example, the exposure risks for most laboratory work in Biosafety
Level 1 and 2 facilities will be direct contact with the agents, or inadvertent contact


Biosafety Manual


November, 2008









exposures through contaminated work environments. Secondary barriers in these
laboratories may include separation of the laboratory work area from public access,
availability of a decontamination facility (e.g., autoclave), and hand washing facilities.

As the risk for aerosol transmission increases, higher levels of primary containment and
multiple secondary barriers may become necessary to prevent infectious agents from
escaping into the environment. Such design features could include specialized
ventilation systems to assure directional air flow, air treatment systems to decontaminate
or remove agents from exhaust air, controlled access zones, airlocks as laboratory
entrances, or separate buildings or modules for isolation of the laboratory. Design
engineers for laboratories may refer to specific ventilation recommendations as found in
the Applications Handbook for Heating, Ventilation, and Air-conditioning (HVAC)
published by the American Society of Heating, Refrigerating, and Air-conditioning
Engineers (ASHRAE).

Biosafety Levels
Four biosafety levels (BSLs) are described which consist of combinations of laboratory
practices and techniques, safety equipment, and laboratory facilities used to control
hazardous biological materials. Each combination is specifically appropriate for the
operations performed, the documented or suspected routes of transmission of the
infectious agents, and for the laboratory function or activity.

The recommended biosafety levels) for the organisms represent those conditions under
which the agent can ordinarily be safely handled. The laboratory director is specifically
and primarily responsible for assessing risks and for appropriately applying the
recommended biosafety levels. Generally, work with known agents should be conducted
at the biosafety level recommended. When specific information is available to suggest
that virulence, pathogenicity, antibiotic resistance patterns, vaccine and treatment
availability, or other factors are significantly altered, more (or less) stringent practices
may be specified.

Biosafety Level 1 practices, safety equipment, and facilities are appropriate for
undergraduate and secondary educational training and teaching laboratories, and for other
facilities in which work is done with defined and characterized strains of viable
microorganisms not known to cause disease in healthy adult humans. Bacillus subtilis,
Naegleria gruberi, and infectious canine hepatitis virus are representative of those
microorganisms meeting these criteria. Many agents not ordinarily associated with
disease processes in humans are, however, opportunistic pathogens and may cause
infection in the young, the aged, and immunodeficient or immunosuppressed individuals.
Vaccine strains which have undergone multiple in vivo passages should not be
considered avirulent simply because they are vaccine strains.

Biosafety Level 1 represents a basic level of containment that relies on standard
microbiological practices with no special primary or secondary barriers recommended,
other than a sink for hand washing.


Biosafety Manual


November, 2008










Biosafety Level 2 practices, equipment, and facilities are applicable to clinical,
diagnostic, teaching, and other facilities in which work is done with the broad spectrum
of indigenous moderate-risk agents present in the community and associated with human
disease of varying severity. With good microbiological techniques, these agents can be
used safely in activities conducted on the open bench, provided the potential for
producing splashes or aerosols is low. Hepatitis B virus, Salmonellae, and Toxoplasma
spp. are representative of microorganisms assigned to this containment level. Biosafety
Level 2 is appropriate when work is done with any human-derived blood, body fluids, or
tissues where the presence of an infectious agent may be unknown. (Laboratory
personnel working with human-derived materials should refer to the Bloodborne
Pathogen Standard for specific, required precautions).

Primary hazards to personnel working with these agents relate to
accidental percutaneous or mucous membrane exposures, or
ingestion of infectious materials. Extreme precaution with
contaminated needles or sharp instruments must be emphasized.
Even though organisms routinely manipulated at BSL-2 are not
known to be transmissible by the aerosol route, procedures with
aerosol or high splash potential that may increase the risk of such personnel exposure
must be conducted in primary containment equipment, or devices such as a BSC or safety
centrifuge cups. Other primary barriers should be used, as appropriate, such as splash
shields, face protection, gowns, and gloves.

Secondary barriers such as hand washing and waste decontamination facilities must be
available to reduce potential environmental contamination.

Biosafety Level 3 practices, safety equipment, and facilities are applicable to clinical,
diagnostic, teaching, research, or production facilities in which work is done with
indigenous or exotic agents with a potential for respiratory transmission, and which may
cause serious and potentially lethal infection. Mycobacterium tuberculosis, St. Louis
encephalitis virus, and Coxiella burnetii are representative of microorganisms assigned to
this level. Primary hazards to personnel working with these agents relate to
autoinoculation, ingestion, and exposure to infectious aerosols.

At Biosafety Level 3, more emphasis is placed on primary and secondary barriers to
protect personnel in contiguous areas, the community, and the environment from
exposure to potentially infectious aerosols. For example, all laboratory manipulations
should be performed in a BSC or other enclosed equipment, such as a gas-tight aerosol
generation chamber. Secondary barriers for this level include controlled access to the
laboratory and a specialized ventilation system that minimizes the release of infectious
aerosols from the laboratory.

Biosafety Level 4 practices, safety equipment, and facilities are applicable for work with
dangerous and exotic agents that pose a high individual risk of life-threatening disease,
which may be transmitted via the aerosol route, and for which there is no available


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vaccine or therapy. Additionally, agents with a close or identical antigenic relationship
to Biosafety Level 4 agents should also be handled at this level. When sufficient data are
obtained, work with these agents may continue at this level or at a lower level. Viruses
such as Marburg or Congo-Crimean hemorrhagic fever are manipulated at Biosafety
Level 4.

The primary hazards to personnel working with Biosafety Level 4 agents are respiratory
exposure to infectious aerosols, mucous membrane exposure to infectious droplets, and
autoinoculation. All manipulations of potentially infectious diagnostic materials,
isolates, and naturally or experimentally infected animals pose a high risk of exposure
and infection to laboratory personnel, the community, and the environment.

The laboratory worker's complete isolation of aerosolized infectious materials is
accomplished primarily by working in a Class III BSC or a full-body, air-supplied
positive-pressure personnel suit. The Biosafety Level 4 facility itself is generally a
separate building or completely isolated zone with complex, specialized ventilation and
waste management systems to prevent release of viable agents to the environment.

The laboratory director is specifically and primarily responsible for the safe operation of
the laboratory. His/her knowledge and judgment are critical in assessing risks and
appropriately applying these recommendations. The recommended biosafety level
represents those conditions under which the agent can ordinarily be safely handled.
Special characteristics of the agents used, the training and experience of personnel, and
the nature or function of the laboratory may further influence the director in applying
these recommendations.

Animal Facilities
Four biosafety levels are also described for activities involving infectious disease work
with experimental mammals. These four combinations of practices, safety equipment,
and facilities are designated Animal Biosafety Levels 1, 2, 3, and 4, and provide
increasing levels of protection to personnel and the environment.

Clinical Laboratories
Clinical laboratories, especially those in health care facilities, receive clinical specimens
with requests for a variety of diagnostic and clinical support services. Typically, the
infectious nature of clinical material is unknown, and specimens are often submitted with
a broad request for microbiological examination for multiple agents (e.g., sputa submitted
for "routine," acid-fast, and fungal cultures). It is the responsibility of the laboratory
director to establish standard procedures in the laboratory that realistically address the
issue of the infectious hazard of clinical specimens.

Except in extraordinary circumstances (e.g., suspected hemorrhagic fever), the initial
processing of clinical specimens and identification of isolates can be done safely at
Biosafety Level 2, the recommended level for work with bloodborne pathogens such as


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hepatitis B virus and HIV. The containment elements described in Biosafety Level 2 are
consistent with the Occupational Exposure to Bloodborne Pathogens Standard from the
Occupational Safety and Health Administration (OSHA), that requires the use of specific
precautions with all clinical specimens of blood or other potentially infectious material
(Universal Precautions). Additionally, other recommendations specific for clinical
laboratories may be obtained from the National Committee for Clinical Laboratory
Standards.

Biosafety Level 2 recommendations and OSHA requirements focus on the prevention of
percutaneous and mucous membrane exposures to clinical material. Primary barriers
such as biological safety cabinets (Class I or II) should be used when performing
procedures that might cause splashing, spraying, or splattering of droplets. Biological
safety cabinets should also be used for the initial processing of clinical specimens when
the nature of the test requested or other information is suggestive that an agent readily
transmissible by infectious aerosols is likely to be present (e.g., M. tuberculosis), or when
the use of a biological safety cabinet (Class II) is indicated to protect the integrity of the
specimen.

The segregation of clinical laboratory functions and limiting or restricting access to such
areas is the responsibility of the laboratory director. It is also the director's responsibility
to establish standard, written procedures that address the potential hazards and the
required precautions to be implemented.

Importation and Interstate Shipment of Certain Biomedical Materials
The importation of etiologic agents and vectors of human diseases is subject to the
requirements of the Public Health Service Foreign Quarantine regulations. Companion
regulations of the Public Health Service and the Department of Transportation specify
packaging, labeling, and shipping requirements for etiologic agents and diagnostic
specimens shipped in interstate commerce.

The U. S. Department of Agriculture regulates the importation and interstate shipment of
animal pathogens and prohibits the importation, possession, or use of certain exotic
animal disease agents that pose a serious disease threat to domestic livestock and poultry.

Please see the section entitled "Shipment of Biological Materials" for more information
on permits.



Biological Safety Levels
The following is from Biosafety in Microbiological and Biomedical Laboratories, 1999, HHS publication
No. (CDC) 93-8395, Centers for Disease Control & Prevention/National Institutes of Health


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Biosafety Level 1
Biosafety Level 1 is suitable for work involving well-characterized agents not known to
cause disease in healthy adult humans, and of minimal potential hazard to laboratory
personnel and the environment. The laboratory is not necessarily separated from the
general traffic patterns in the building. Work is generally conducted on open bench tops
using standard microbiological practices. Special containment equipment or facility
design is not required nor generally used. Laboratory personnel have specific training in
the procedures conducted in the laboratory and are supervised by a scientist with general
training in microbiology or a related science.

The following standard and special practices, safety equipment, and facilities apply to the
handling of agents assigned to Biosafety Level 1:

A. Standard Microbiological Practices
1. Access to the laboratory is limited or restricted at the discretion of the
laboratory director when experiments or work with cultures and specimens
are in progress.
2. Persons wash their hands after they handle viable
materials and animals, after removing gloves, and
before leaving the laboratory.
3. Eating, drinking, use of tobacco products, handling
contact lenses, and applying cosmetics are not
permitted in the work areas where there is
reasonable likelihood of exposure to potentially infectious materials.
Persons who wear contact lenses in laboratories should also wear goggles
or a face shield. Food is stored outside the work area in cabinets or
refrigerators designated and used for that purpose only.
4. Mouth pipetting is prohibited; mechanical pipetting devices are used.
5. All procedures are performed carefully to minimize the creation of
splashes or aerosols.
6. Work surfaces are decontaminated at least once a day and after any spill of
viable material.
7. All cultures, stocks, and regulated wastes are decontaminated before
disposal by an approved decontamination method, such as autoclaving.
Materials to be decontaminated outside of the immediate laboratory are to
be placed in a durable, leak-proof container and closed for transport from
the laboratory. Materials to be decontaminated off-site from the
laboratory are packaged in accordance with applicable local, state, and
federal regulations, before removal from the facility.
8. An insect and rodent control program is in effect.

B. Special Practices: None

C. Safety Equipment (Primary Barriers)


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1. Special containment devices or equipment such as a biological safety
cabinet are generally not required for manipulations of agents assigned to
Biosafety Level 1.
2. It is recommended that laboratory coats, gowns, or uniforms are worn to
prevent contamination or soiling of street clothes.
3. Gloves should be worn if the skin on the hands is broken or if a rash
exists.
4. Protective eyewear should be worn for anticipated splashes of
microorganisms or other hazardous materials to the face.

D. Laboratory Facilities (Secondary Barriers)
1. Each laboratory contains a sink for hand washing
2. The laboratory is designed so that it can be easily cleaned. Walls and
floors must be constructed of water impervious materials that will stand up
to hard disinfectants. Carpeting is not allowed in laboratories.
3. Bench tops are impervious to water and resistant to acids, alkalis, organic
solvents, and moderate heat.
4. Laboratory furniture is sturdy and should not be cloth upholstered. Spaces
between benches, cabinets, and equipment are accessible for cleaning.
5. If the laboratory has windows that open, they are fitted with fly screens.

Biosafety Level 2
Biosafety Level 2 is similar to Level 1 and is suitable for work involving agents of
moderate potential hazard to personnel and the environment. It differs in four ways.

1. Laboratory personnel must have specific training in handling pathogenic
agents and should be directed by competent scientists.
2. Access to the laboratory must be limited when work is being conducted.
3. Extreme precautions must be taken with contaminated sharp items.
4. Certain procedures in which infectious aerosols or splashes may be created
must be conducted in biological safety cabinets or other physical containment
equipment.

The following standard and special practices, safety equipment, and facilities apply to the
use of agents assigned to Biosafety Level 2:

A. Standard Microbiological Practices
1. Access to the laboratory is limited or restricted at the discretion of the
laboratory director when experiments are in progress.
2. Persons wash their hands after they handle viable materials and animals,
after removing gloves, and before leaving the laboratory.
3. Eating, drinking, the use of tobacco products, handling contact lenses, and
applying cosmetics are not permitted in the work areas. Persons who wear
contact lenses in laboratories should also wear goggles or a face shield.


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Food is stored outside the work area in cabinets or refrigerators designated
for that purpose only.
4. Mouth pipetting is prohibited; mechanical pipetting devices are used.
5. All procedures are performed carefully to minimize the creation of
splashes or aerosols.
6. Work surfaces are decontaminated at least once a day and after any spill of
viable material.
7. All cultures, stocks, and regulated wastes are decontaminated before
disposal by an approved decontamination method, such as autoclaving.
Materials to be decontaminated outside of the immediate laboratory are to
be placed in a durable, leak-proof container and closed for transport from
the laboratory. Materials to be decontaminated off-site from the
laboratory are packaged in accordance with applicable local, state, and
federal regulations, before removal from the facility.
8. An insect and rodent control program is in effect.

B. Special Practices
1. Access to the laboratory is limited or restricted by the laboratory director
when work with infectious agents is in progress. In general, persons who
are at increased risk of acquiring infection or for whom infection may be
unusually hazardous are not allowed in the laboratory or animal rooms.
For example, persons who are immunocompromised or
immunosuppressed may be at risk of acquiring infections. The laboratory
director has the final responsibility for assessing each circumstance and
determining who may enter or work in the laboratory.
2. The laboratory director establishes policies and procedures whereby only
persons who have been advised of the potential hazard and meet specific
entry requirements (e.g., immunization) enter the laboratory or animal
rooms.
3. When the infectious agents) in use in the laboratory require special
provisions or entry (e.g., immunization), a hazard warning sign
incorporating the universal biohazard symbol is posted on the access door
to the laboratory work area. The hazard warning sign identifies the
infectious agent, lists the name and telephone number of the laboratory
director or other responsible personss, and indicates the special
requirements) for entering the laboratory.
4. Laboratory personnel receive appropriate immunizations or tests for the
agents handled or potentially present in the laboratory (e.g., hepatitis B
vaccine or TB skin testing).
5. When appropriate, considering the agents) handled, baseline serum samples for
laboratory and other at-risk personnel are collected and stored. Additional serum
specimens may be collected periodically, depending on the agents handled or the
function of the facility.
6. A Biosafety manual is prepared or adopted. Personnel are advised of special
hazards and are required to read and to follow instructions on practices and
procedures.


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7. Laboratory personnel receive appropriate training on the potential hazards
associated with the work involved, the necessary precautions to prevent
exposures, and the exposure evaluation procedures. Personnel receive annual
updates, or additional training as necessary for procedural or policy changes.
8. A high degree of precaution must always be taken with any contaminated sharp
items, including needles and syringes, slides, pipettes, capillary tubes, and
scalpels. Needles and syringes or other sharp instruments should be restricted in
the laboratory for use only when there is no alternative, such as parenteral
injection, phlebotomy, or aspiration of fluids from laboratory animals and
diaphragm bottles. Plasticware should be substituted for glassware whenever
possible.
a. Only needle-locking syringes or disposable syringe-needle units (i.e.,
needle is integral to the syringe) are used for injection or aspiration of
infectious materials. Used disposable needles must not be bent, sheared,
broken, recapped, removed from disposable syringes, or otherwise
manipulated by hand before disposal; rather, they must be carefully placed
in conveniently located puncture-resistant containers used for sharps
disposal. Non-disposable sharps must be placed in a hard-walled
container for transport to a processing area for decontamination,
preferable by autoclaving.
b. Syringes that re-sheathe the needle, needle-less systems, and other safe
devices should be used when appropriate.
c. Broken glassware must not be handled directly by hand, but must be
removed by mechanical means such as a brush and dustpan, tongs, or
forceps. Containers of contaminated needles, sharp equipment, and
broken glass are decontaminated before disposal, according to any local,
state, or federal regulations.
9. Cultures, tissues, or specimens of body fluids are placed in a container that
prevents leakage during collection, handling, processing, storage, transport, or
shipping.
10. Laboratory equipment and work surfaces should be decontaminated with an
appropriate disinfectant on a routine basis, after work with infectious materials is
finished, and especially after overt spills, splashes, or other contamination by
infectious materials. Contaminated equipment must be decontaminated according
to any local, state, or federal regulations before it is sent for repair or
maintenance. Equipment must also be decontaminated before removal from the
facility when it must be packaged for transport. Packaging and shipment shall be
in accordance with applicable local, state, or federal regulations,.
11. Spills and accidents that result in overt exposures to infectious materials are
immediately reported to the laboratory director. Medical evaluation, surveillance,
and treatment are provided as appropriate and written records are maintained.
12. Animals not involved in work being performed aren't permitted in the lab.

C. Safety Equipment (Primary Barriers)


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1. Properly maintained biological safety cabinets, preferably Class II, or
other appropriate personal protective equipment or physical containment
devices are used under the following conditions.
a. Use Biological Safety Cabinets whenever procedures with a
potential for creating infectious aerosols or splashes are conducted.
These may include centrifuging, grinding, blending, vigorous
shaking or mixing, sonic disruption, opening containers of
infectious materials whose internal pressures may be different
from ambient pressures, inoculating animals intranasally, and
harvesting infected tissues from animals or eggs.
b. Use Biological Safety Cabinets whenever high concentrations or
large volumes of infectious agents are used. Such materials may
be centrifuged in the open laboratory if sealed rotor heads or
centrifuge safety cups are used, and if these rotors or safety cups
are opened only in a biological safety cabinet.
2. Face protection (goggles, mask, face shield or other splatter guards) is
required to prevent splashes or sprays of infectious or other hazardous
materials to the face, when the microorganisms must be manipulated
outside the BSC.
3. Protective laboratory coats, gowns, smocks, or uniforms designated for lab
use are worn while in the laboratory. This protective clothing is removed
and left in the laboratory before leaving for non-laboratory areas (e.g.,
cafeteria, library, and administrative offices). All protective clothing is
either disposed of in the laboratory or laundered by the institution;
personnel should never take it home.
4. Gloves are required when handling infected animals and when hands may
contact infectious materials, contaminated surfaces or equipment.
Wearing two pairs of gloves may be appropriate; if a spill or splatter
occurs, the hand will be protected after the contaminated glove is
removed. Gloves are disposed of when contaminated, removed when
work with infectious materials is completed, and are never worn outside
the laboratory. Disposable gloves are not washed or reused.

D. Laboratory Facilities (Secondary Barriers)
1. Each laboratory contains a sink for hand washing.
2. The laboratory is designed so that it can be easily cleaned. Walls and
floors must be constructed of water impervious material that will stand up
to harsh disinfectants. Carpeting is not allowed in laboratory facilities.
3. Bench tops are impervious to water and resistant to acids, alkalis, organic
solvents, and moderate heat.
4. Laboratory furniture is sturdy, and spaces between benches, cabinets, and
equipment are accessible for cleaning. No cloth or fabric seating is
permitted.
5. If the laboratory has windows that open, they are fitted with fly screens.


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6. A method for decontamination of infectious or regulated laboratory wastes
is available (e.g., autoclave, chemical disinfection, incinerator, or other
approved decontamination system).
7. An eyewash/safety shower facility is readily available.
8. All laboratories require single pass air that is not then recirculated to any
other area of the facility. Laboratories should be negative pressure to
surrounding areas to prevent accidental spread of potentially infectious or
recombinant agents.
9. Install biological safety cabinets in such a manner that fluctuations of the
room supply and exhaust air do not cause the biological safety cabinets to
operate outside their parameters for containment. Locate biological safety
cabinets away from doors, from windows that can be opened, from heavily
traveled laboratory areas, and from other potentially disruptive equipment
so as to maintain the biological safety cabinets' air flow parameters for
containment.

Biosafety Level 3
Biosafety Level 3 is applicable to clinical, diagnostic, teaching, research, or production
facilities in which work is done with indigenous or exotic agents which may cause
serious or potentially lethal disease as a result of exposure by the inhalation route.
Laboratory personnel have specific training in handling pathogenic and potentially lethal
agents, and are supervised by competent scientists who are experienced in working with
these agents.

All procedures involving the manipulation of infectious materials are conducted within
biological safety cabinets or other physical containment devices, or by personnel wearing
appropriate personal protective clothing and equipment. The laboratory has special
engineering and design features.

It is recognized, however, that many existing facilities may not have all the facility
safeguards recommended for Biosafety Level 3 (e.g., access zone, sealed penetrations,
directional airflow, etc.). In these circumstances, acceptable safety may be achieved for
routine or repetitive operations (e.g. diagnostic procedures involving the propagation of
an agent for identification, typing, and susceptibility testing) in Biosafety Level 2
facilities. However, the recommended Standard Microbiological Practices, Special
Practices, and Safety Equipment for Biosafety Level 3 must be rigorously followed. The
decision to implement this modification of Biosafety Level 3 recommendations should be
made only by the lab director. It is strongly recommended that persons have a minimum
of 120 hour work experience in a Level 2 laboratory to qualify to work at Level 3.

The following standard and special safety practices, equipment, and facilities apply to the
use of agents assigned to Biosafety Level 3:


A. Standard Microbiological Practices


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1. Access to the laboratory is limited or restricted at the discretion of the
laboratory director when experiments are in progress.
2. Persons wash their hands after handling infectious materials and animals,
after removing gloves, and when they leave the laboratory.
3. Eating, drinking, the use of tobacco products, handling contact lenses and
applying cosmetics or lip balm are not permitted in the laboratory.
Persons who wear contact lenses in laboratories should also wear goggles
or a face shield. Food is stored outside the work area in cabinets or
refrigerators designated for this purpose only.
4. Mouth pipetting is prohibited; mechanical pipetting devices are used.
5. Policies for the safe handling of sharps are instituted.
6. All procedures are performed carefully to minimize the creation of
aerosols.
7. Work surfaces are decontaminated at least once a day and immediately
after any spill of viable material.
8. All cultures, stocks, and regulated wastes are decontaminated before
disposal by an approved decontamination method, such as autoclaving.
Materials to be decontaminated outside of the immediate laboratory are to
be placed in a durable, leak-proof container and closed for transport from
the laboratory. Infectious wastes from BSL-3 laboratories must be
decontaminated before removal for off-site disposal.
9. An insect and rodent control program is in effect.
10. Policies for the safe handling of sharps are instituted.

B. Special Practices
1. Laboratory doors are kept closed at all times.
2. The laboratory director controls access to the laboratory and restricts
access to persons whose presence is required for program or support
purposes. For example, persons who are immunocompromised or
immunosuppressed, or for whom infection may be unusually hazardous,
are not allowed in the laboratory or animal rooms. The director has the
final responsibility for assessing each circumstance and determining who
may enter or work in the laboratory. No minors are allowed in level 3
laboratories.
3. The laboratory director establishes policies and procedures whereby only
persons who have been advised of the potential biohazard, who meet any
specific entry requirements (e.g., immunizations), and who comply with
all entry and exit procedures, enter the laboratory or animal rooms.
4. When infectious materials or infected animals are present in the laboratory
or containment module, a hazard warning sign incorporating the universal
biohazard symbol, is posted on all laboratory and animal room access
doors. The hazard warning sign identifies the agent, lists the name and
telephone number of the laboratory director or other responsible personss,
and indicates any special requirements for entering the laboratory, such as
the need for immunizations, respirators, or other personal protective
measures.


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5. Laboratory personnel receive the appropriate immunizations or tests for
the agents handled or potentially present in the laboratory (e.g., hepatitis B
vaccine or TB skin testing and periodic testing.)
6. Baseline serum samples are collected and stored for all laboratory and
other at-risk personnel. Additional serum specimens may be collected
periodically, depending on the agents handled or the function of the
laboratory.
7. A Biosafety Manual specific to the laboratory is prepared or adopted.
Personnel are advised of special hazards and are required to read and to
follow instructions on practices and procedures. An SOP manual is
prepared to cover all procedures and activities.
8. Laboratory personnel receive appropriate training on the potential hazards
associated with the work involved, the necessary precautions to prevent
exposures, and the exposure evaluation procedures. Personnel receive
annual updates, or additional training as necessary for procedural changes.
9. The laboratory director is responsible for insuring that all personnel
demonstrate proficiency in standard microbiological practices and
techniques, and in the practice and operations specific to the laboratory
facility, before working with organisms at Biosafety Level 3. This might
include prior experience in handling human pathogens or cell cultures, or
a specific training program provided by the laboratory director or other
competent scientist proficient in safe microbiological practices and
techniques.
10. A high degree of precaution must always be taken with all contaminated
sharp items, including needles and syringes, slides, pipettes, capillary
tubes, and scalpels. Needles and syringes or other sharp instruments
should be restricted in the laboratory for use only when there is no
alternative, such as parenteral injection, phlebotomy, or aspiration of
fluids from laboratory animal and diaphragm bottles. Plasticware should
be substituted for glassware whenever possible.
a. Only needle-locking syringes or disposable syringe needle units
(i.e., needle is integral to the syringe) are used for injection or
aspiration of infectious materials. Used disposable needles must
not be bent, sheared, broken, recapped, removed from disposable
syringes, or otherwise manipulated by hand before disposal; rather,
they must be carefully placed in conveniently located puncture-
resistant containers used for sharps disposal. Non-disposable
sharps must be placed in a hard-walled container for transport to a
processing area for decontamination, preferably by autoclaving.
b. Syringes that re-sheathe the needle, needle-less systems, and other
safe devices should be used when appropriate.
c. Broken glassware must not be handled directly by hand, but must
be removed by mechanical means such as a brush and dustpan,
tongs, or forceps. Containers of contaminated needles, sharp
equipment, and broken glass should be decontaminated before
disposal, in accordance with any local, state, or federal regulations.


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11. All manipulations involving infectious materials are conducted in
biological safety cabinets or other physical containment devices within the
containment module. No work in open vessels is conducted on the open
bench.
12. Laboratory equipment and work surfaces should be decontaminated with
an appropriate disinfectant on a routine basis, after work with infectious
materials is finished. It must be cleaned and decontaminated after overt
spills, splashes, or other contamination with infectious materials.
Contaminated equipment should also be decontaminated before it is sent
for repair or maintenance. In addition, it must be packaged for transport in
accordance with applicable local, state, or federal regulations, before
removal from the facility. Plastic-backed paper toweling can be used on
non-perforated work surfaces within biological safety cabinets to facilitate
clean up.
13. Cultures, tissues, or specimens of body fluids are placed in a container that
prevents leakage during collection, handling, processing, storage,
transport, or shipping.
14. All potentially contaminated materials (e.g., gloves, lab coats, etc.) from
laboratories or animal rooms are decontaminated before disposal or reuse.
15. Spills of infectious materials are decontaminated, contained and cleaned
up by appropriate professional staff, or others properly trained and
equipped to work with concentrated infectious material.
16. Spills and accidents that result in overt or potential exposures to infectious
materials are immediately reported to the laboratory director. Appropriate
medical evaluation, surveillance, and treatment are provided and written
records are maintained.
17. Animals and plants not related to the work being conducted are not
permitted in the laboratory.

C. Safety Equipment (Primary Barriers)
1. Properly maintained biological safety cabinets are used (Class II or III) for
all manipulation of infectious materials. Biological Safety Cabinets (BSC)
should be located away from doors, air supplies and other heavily traveled
areas. Biosafety Cabinets must be certified at least yearly.
2. Outside of a BSC, appropriate combinations of personal protective
equipment are used (e.g., special protective clothing, masks, gloves, face
protection, or respirators) in combination with physical containment
devices (e.g., centrifuge safety cups, sealed centrifuge rotors, or
containment caging for animals).
3. Biological Safety Cabinets must be used for manipulations of cultures and
clinical or environmental materials that may be a source of infectious
aerosols. The aerosol challenge of experimental animals; harvesting of
tissues or fluids from infected animals, and embryonated eggs, and
necropsy of infected animals also require the use of BSCs.


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4. Face protection (goggles and mask, or face shield) is worn for
manipulations of infectious materials outside of a biological safety
cabinet.
5. Respiratory protection is worn when aerosols cannot be safety contained
(e.g., outside of a biological safety cabinet), and in rooms containing
infected animals.
6. Protective laboratory clothing such as solid-front or wrap-around gowns,
scrub suits, or coveralls must be worn in, and not worn outside, the
laboratory. Reusable laboratory clothing is to be decontaminated before
being laundered. Change protective clothing immediately if
contaminated.
7. Gloves must be worn when handling infected animals and when hands
may contact infectious materials and contaminated surfaces or equipment.
Disposable gloves should be discarded when contaminated, and never
washed for reuse. Always wash hands between glove changes.

D. Laboratory Facilities (Secondary Barriers)
1. The laboratory is separated from areas that are open to unrestricted traffic
flow within the building. Passage through two sets of self-closing doors is
the basic requirement for entry into the laboratory from access corridors or
other contiguous areas. A clothes change room (shower optional) may be
included in the passageway.
2. Each laboratory contains a sink for hand washing. The sink is foot, elbow,
or automatically operated and is located near the laboratory exit door.
3. The interior surfaces of walls, floors, and ceilings are water-resistant so
that they can be easily cleaned. Penetrations in these surfaces are sealed
or capable of being sealed to facilitate decontamination. Floors should be
monolithic and coved to the walls.
4. Bench tops are impervious to water and resistant to acids, alkalis, organic
solvents, and moderate heat.
5. Laboratory furniture is sturdy, and spaces between benches, cabinets, and
equipment are accessible for cleaning. No fabric materials are allowed.
6. Windows in the laboratory are closed and sealed.
7. A method for decontaminating all laboratory wastes is available,
preferably within the laboratory (i.e., autoclave, chemical disinfection,
incineration, or other approved decontamination method).
8. A ducted exhaust air ventilation system is provided. This system creates
directional airflow that draws air from "clean" areas into the laboratory
toward "contaminated" areas. The exhaust air is not recirculated to any
other area of the building, and is discharged to the outside through a
HEPA filtration system. The outside exhaust must be dispersed away
from occupied areas and air intakes. Laboratory personnel must verify
that the direction of the airflow (into the laboratory) is proper. Visual
monitoring devices are recommended.
9. The High Efficiency Particulate Air (HEPA)-filtered exhaust air from
Class II or Class III biological safety cabinets is discharged directly to the


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outside or through the building exhaust system. If the HEPA-filtered
exhaust air from Class II or III biological safety cabinets is to be
discharged to the outside through the building exhaust air system, it is
connected to this system in a manner (e.g., thimble unit connection) that
avoids any interference with the air balance of the cabinets or building
exhaust system. Exhaust air from Class II biological safety cabinets may
be recirculated within the laboratory if the cabinet is tested and certified at
least every twelve months.
10. Continuous flow centrifuges or other equipment that may produce aerosols
are contained in devices that exhaust air through HEPA filters before
discharge into the laboratory.
11. Vacuum lines are protected with liquid disinfectant traps and HEPA
filters, or their equivalent, which are routinely maintained and replaced as
needed.
12. An eyewash/safety shower is readily available.
13. A Biosafety Level 3 facility design and operational procedures must be
documented. The facility must be tested for verification that the design
and operational parameters have been met prior to operation. Facilities
should be re-verified/certified at least annually (HEPAs and air systems).
14. Additional environmental protection (e.g. personnel showers, containment
of other piped services and the provision for effluent decontamination)
should be added when recommended by the agent summary statement, as
determined by risk assessment, the site conditions or other applicable
federal, state, or local regulations.
15. Level 3 laboratories and animal areas are audited by EH&S quarterly.
16. Openings such as around ducts, spaces between doors and frames and
other penetrations must be capable of being sealed to facilitate
decontamination.


Animal Biosafety Level 1 (ABSL-1)

Animal Biosafety Level 1 (ABSL-1) is suitable
for work involving well characterized agents
that are not known to cause disease in healthy
adult humans, and that are of minimal potential
hazard to laboratory personnel and the
environment.



A. Standard Practices
1. The animal facility director establishes polices, procedures, and protocols
for emergency situations. Each project is subject to pre-approval by the
Institutional Animal Care and Use Committee (IACUC) and the


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Institutional Biosafety Committee). Any special practices are approved at
this time.
2. Only those persons required for program or support purposes are
authorized to enter the facility. Before entering, persons are advised of the
potential biohazards and are instructed on the appropriate safeguards.
3. An appropriate medical surveillance program is in place.
4. A safety manual is prepared or adopted. Personnel are advised of special
hazards, and are required to read and follow instructions on practices and
procedures.
5. Eating, drinking, smoking, handling contact lenses, applying cosmetics,
and storing food for human use should only be done in designated areas
and are not permitted in animal or procedure rooms.
6. All procedures are carefully performed to minimize the creation of
aerosols or splatters.
7. Work surfaces are decontaminated after use or after any spill of viable
materials.
8. All wastes from the animal room (including animal tissues, carcasses, and
contaminated bedding) are transported from the animal room in leak-
proof, covered containers for appropriate disposal in compliance with
applicable institutional or local requirements. Incineration is
recommended.
9. Policies for the safe handling of sharps are instituted.
10. Personnel wash their hands after handling cultures and animals, after
removing gloves, and before leaving the animal facility.
11. A biohazard sign must be posted on the entrance to the animal room
whenever infectious agents are present. The hazard warning sign identifies
the infectious agents) in use, lists the name and telephone number of the
responsible personss, and indicates the special requirements for entering
the animal room (e.g., the need for immunizations and respirators).
12. An insect and rodent control program is in effect.
B. Special Practices: None
C. Safety Equipment (Primary Barriers):
1. The wearing of laboratory coats, gowns, and/or uniforms in the facility is
recommended. Laboratory coats remain in the animal room. Gowns and
uniforms are not worn outside the facility.
2. Persons having contact with non-human primates should assess their risk
of mucous membrane exposure and wear appropriate eye and face
protection.


D. Facilities (Secondary Barriers)
1. The animal facility is separated from areas that are open to unrestricted
personnel traffic within the building.
2. External facility doors are self-closing and self-locking. Doors to animal
rooms open inward, are self-closing, and are kept closed when


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experimental animals are present. Cubicle room inner doors may open
outward or be horizontal or vertical sliding.
3. The animal facility is designed, constructed, and maintained to facilitate
cleaning and housekeeping. The interior surfaces (walls, floors, and
ceilings) are water resistant.
4. Internal facility appurtenances, such as light fixtures, air ducts, and utility
pipes, are arranged to minimize horizontal surface areas.
5. Windows are not recommended. Any windows must be resistant to
breakage. Where possible, windows should be sealed. If the animal facility
has windows that open, they are fitted with fly screens.
6. If floor drains are provided, the traps are always filled with water and/or
an appropriate disinfectant.
7. Ventilation should be provided in accordance with the Guide for Care and
Use ofLaboratory Animals, latest edition. No recirculation of exhaust air
should occur. It is recommended that animal rooms maintain negative
pressure compared to adjoining hallways.
8. The facility has a hand washing sink.
9. Cages are washed manually or in a cage washer. The mechanical cage
washer should have a final rinse temperature of at least 180F.
10. Illumination is adequate for all activities, avoiding reflections and glare
that could impede vision.

Animal Biosafety Level 2 (ABSL-2)


1Animal Biosafety Level 2 involves practices for work
with those agents associated with human disease. It
addresses hazards from ingestion as well as from
percutaneous and mucous membrane exposure. ABSL-2
builds upon the practices, procedures, containment
equipment, and facility requirements of ABSL-1.



A. Standard Practices
1. Aside from the standard policies, procedures, and protocols for emergency
situations established by the facility director, appropriate special policies
and procedures should be developed as needed and approved by the
Institutional Animal Care and Use Committee (IACUC) and the
Institutional Biosafety Committee (IBC).
2. Access to the animal room is limited to the fewest number of individuals
possible. Personnel who must enter the room for program or service
purposes when work is in progress are advised of the potential hazard.
3. An appropriate medical surveillance program is in place. All personnel
receive appropriate immunizations or tests for the agents handled or


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potentially present (e.g., hepatitis B vaccine, TB skin testing). When
appropriate, a serum surveillance system should be implemented.
4. A biosafety manual is prepared or adopted. Personnel are advised of
special hazards, and are required to read and follow instructions on
practices and procedures.
5. Eating, drinking, smoking, handling contact lenses, applying cosmetics,
and storing food for human use should only be done in designated areas
and are not permitted in animal or procedure rooms.
6. All procedures are carefully performed to minimize the creation of
aerosols or splatters.
7. Equipment and work surfaces in the room are routinely decontaminated
with an effective disinfectant after work with the infectious agent, and
especially after overt spills, splashes, or other contamination by infectious
materials.
8. All infectious samples are collected, labeled, transported, and processed in
a manner that contains and prevents transmission of the agentss. All
wastes from the animal room (including animal tissues, carcasses,
contaminated bedding, unused feed, sharps, and other refuse) are
transported from the animal room in leak-proof, covered containers for
appropriate disposal in compliance with applicable institutional or local
requirements. The outer surface of the containers is disinfected prior to
moving the material. Autoclaving of the contents prior to incineration is
recommended.
9. Policies for the safe handling of sharps are instituted:
a. Needles and syringes or other sharp instruments are
restricted for use in the animal facility only when there is
no alternative, such as for parenteral injection, blood
collection, or aspiration of fluids from laboratory animals
and diaphragm bottles.
b. Syringes that re-sheathe the needle, needle-less systems,
and other safe devices should be used when appropriate.
c. Plasticware should be substituted for glassware whenever
possible.
10. Personnel wash their hands after handling cultures and animals, after
removing gloves, and before leaving the animal facility.
11. A biohazard sign must be posted on the entrance to the animal room
whenever infectious agents are present. The hazard warning sign identifies
the infectious agents) in use, lists the name and telephone number of the
responsible personss, and indicates the special requirements (e.g., the
need for immunizations and respirators) for entering the animal room.
12. An insect and rodent control program is in effect.

B. Special Practices
1. Animal care laboratory and support personnel receive appropriate training
on the potential hazards associated with the work involved, the necessary
precautions to prevent exposures, and the exposure evaluation procedures.


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Personnel receive annual updates, or additional training as necessary for
procedural or policy changes. Records of all training provided are
maintained. In general, persons who may be at increased risk of acquiring
infection, or for whom infection might be unusually hazardous, are not
allowed in the animal facility unless special procedures can eliminate the
extra risk.
2. Only animals used for the experiments) are allowed in the room.
3. All equipment must be appropriately decontaminated prior to removal
from the room.
4. Spills and accidents which result in overt exposures to infectious materials
must be immediately reported to the facility director. Medical evaluation,
surveillance, and treatment are provided as appropriate and written records
are maintained.

C. Safety Equipment (Primary Barriers)
1. Gowns, uniforms, or laboratory coats are worn while in the animal room.
The laboratory coat is removed and left in the animal room. Gowns,
uniforms, and laboratory coats are removed before leaving the animal
facility. Gloves are worn when handling infected animals and when skin
contact with infectious materials is unavoidable
2. Personal protective equipment is used based on risk assessment
determinations. Appropriate face/eye and respiratory protection is worn by
all personnel entering animal rooms that house nonhuman primates.
3. Biological safety cabinets, other physical containment devices, and/or
personal protective equipment (e.g., respirators, face shields) are used
whenever conducting procedures with a high potential for creating
aerosols. These include necropsy of infected animals, harvesting of tissues
or fluids from infected animals or eggs, or intranasal inoculation of
animals.
4. When needed, animals are housed in primary biosafety containment
equipment appropriate for the animal species. Filter top cages are always
handled in properly designed and operating animal bio-containment
cabinets recommended for rodents.

D. Facilities (Secondary Barriers)
1. The animal facility is separated from areas that are open to unrestricted
personnel traffic within the building.
2. Access to the facility is limited by secure locked doors. External doors are
self-closing and self-locking. Doors to animal rooms open inward, are
self-closing, and are kept closed when experimental animals are present.
Cubicle room inner doors may open outward or be horizontal or vertical
sliding.
3. The animal facility is designed, constructed, and maintained to facilitate
cleaning and housekeeping. The interior surfaces (walls, floors, and
ceilings) are water resistant.


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4. Internal facility appurtenances, such as light fixtures, air ducts, and utility
pipes, are arranged to minimize horizontal surface areas.
5. Any windows must be resistant to breakage. Where possible, windows
should be sealed. If the animal facility has windows that open, they are
fitted with fly screens.
6. If floor drains are provided, the traps are always filled with an appropriate
disinfectant.
7. Exhaust air is discharged to the outside without being recirculated to other
rooms. Ventilation should be provided in accordance with criteria from
Guide for Care and Use of Laboratory Animals, latest edition. The
direction of airflow in the animal facility is inward; animal rooms should
maintain negative pressure compared to adjoining hallways.
8. Cages are washed manually or in an appropriate cage washer. The
mechanical cage washer should have a final rinse temperature of at least
180F.
9. An autoclave is available in the animal facility to decontaminate infectious
waste.
10. A hand washing sink is in the animal room where infected animals are
housed, as well as elsewhere in the facility.
11. Illumination is adequate for all activities, avoiding reflections and glare
that could impede vision.

Animal Biosafety Level 3 (ABSL-3)
Animal Biosafety Level 3 involves practices suitable for work with animals infected with
indigenous or exotic agents that present the potential of aerosol transmission and of
causing serious or potentially lethal disease. ABSL-3 builds upon the standard practices,
procedures, containment equipment, and facility requirements of ABSL-2.

A. Standard Practices
1. Aside from the standard policies, procedures, and protocols for emergency
situations established by the facility director, appropriate special policies
and procedures should be developed as needed and approved by the
Institutional Animal Care and Use Committee (IACUC) and the
Institutional Biosafety Committee (IBC).
2. The laboratory or animal facility director limits access to the animal room
to the fewest number of individuals possible. Personnel who must enter
the room for program or service purposes when work is in progress are
advised of the potential hazard.
3. An appropriate medical surveillance program is in place. All personnel
receive appropriate immunizations or tests for the agents handled or
potentially present (e.g., hepatitis B vaccine, TB skin testing). When
appropriate, a serum surveillance system should be implemented. In
general, persons who may be at increased risk of acquiring infection, or
for whom infection might have serious consequences, are not allowed in


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the animal facility unless special procedures can eliminate the extra risk.
Assessment should be made by the occupational health physician.
4. A biosafety manual is prepared or adopted. Personnel are advised of
special hazards, and are required to read and follow instructions on
practices and procedures.
5. Eating, drinking, smoking, handling contact lenses, applying cosmetics,
and storing food for human use should be done only in designated areas
and are not permitted in animal or procedure rooms.
6. All procedures are carefully performed to minimize the creation of
aerosols or splatters.
7. Equipment and work surfaces in the room are routinely decontaminated
with an effective disinfectant after work with the infectious agent, and
especially after overt spills, splashes, or other contamination by infectious
materials.
8. All wastes from the animal room (including animal tissues, carcasses,
contaminated bedding, unused feed, sharps, and other refuse animal
tissues) are transported from the animal room in leak-proof, covered
containers for appropriate disposal in compliance with applicable
institutional or local requirements. Incineration is recommended. The
outer surface of the containers is disinfected prior to moving the material
(see Special Practices #3 below).
9. Policies for the safe handling of sharps are instituted:
a. Needles and syringes or other sharp instruments are restricted in
the animal facility for use only when there is no alternative, such
as for parenteral injection, blood collection, or aspiration of fluids
from laboratory animals and diaphragm bottles.
b. Syringes that re-sheathe the needle, needle-less systems, and other
safety devices should be used when appropriate.
c. Plasticware should be substituted for glassware whenever
possible.
10. Personnel wash their hands after handling cultures and animals, after
removing gloves, and before leaving the animal facility.
11. A biohazard sign must be posted on the entrance to the animal room
whenever infectious agents are present. The hazard warning sign identifies
the infectious agents) in use, lists the name and telephone number of the
responsible personss, and indicates the special requirements for entering
the animal room (e.g., the need for immunizations and respirators).
12. All infectious samples are collected, labeled, transported, and processed in
a manner that contains and prevents transmission of the agentss.
13. Laboratory and support personnel receive appropriate training on the
potential hazards associated with the work involved, the necessary
precautions to prevent exposures, and the exposure evaluation procedures.
As necessary, personnel receive updates and/or additional training on
procedural or policy changes. Records of all training provided are
maintained.
14. An insect and rodent control program is in effect.


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B. Special Practices
1. Cages are autoclaved or thoroughly decontaminated before bedding is
removed and before they are cleaned and washed. Equipment must be
decontaminated according to any local, state, or federal regulations before
being packaged for transport or removal from the facility for repair or
maintenance.
2. A spill procedure is developed and posted. Only personnel properly
trained and equipped to work with infectious materials are to clean up
spills. Spills and accidents that result in overt exposures to infectious
materials must be immediately reported to the facility director. Medical
evaluation, surveillance, and treatment are provided as appropriate and
written records are maintained.
3. All wastes from the animal room must be autoclaved prior to incineration
or other appropriate terminal treatment.
4. Materials not related to the experiment (e.g., plants, animals) are not
permitted in the animal room.

C. Safety Equipment (Primary Barriers)
1. Uniforms or scrub suits are worn by personnel entering
the animal room. Wrap-around or solid-front gowns i
should be worn over this clothing. Front-button laboratory
coats are unsuitable. The gown must be removed and left
in the animal room. Before leaving the animal facility,
scrub suits and uniforms are removed and appropriately
contained and decontaminated prior to laundering or
disposal.
2. Personal protective equipment used is based on risk
assessment determinations.
a. Personal protective equipment is used for all activities involving
manipulations of infectious material or infected animals.
b. Personnel wear gloves when handling infected animals. Gloves are
removed aseptically and autoclaved with other animal room wastes
before disposal.
c. Appropriate face/eye and respiratory protection (e.g., respirators
and face shields) is worn by all personnel entering animal rooms.
d. Boots, shoe covers, or other protective footwear, and disinfectant
foot baths are available and used where indicated.
3. The risk of infectious aerosols from infected animals or their bedding also
can be reduced if animals are housed in containment caging systems, such
as open cages placed in inward flow ventilated enclosures (e.g., laminar
flow cabinets), solid wall and bottom cages covered with filter bonnets, or
other equivalent primary containment systems.
4. Biological safety cabinets and other physical containment devices are used
whenever conducting procedures with a potential for creating aerosols.


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These include necropsy of infected animals, harvesting of tissues or fluids
from infected animals or eggs, or intranasal inoculation of animals. At
BSL-3, all work should be done in a primary barrier; otherwise respirators
should be worn by personnel in the room.

D. Facilities (Secondary Barriers)
1. The animal facility is separated from areas that are open to unrestricted
personnel traffic within the building.
2. Access to the facility is limited by a self-closing and self-locking door.
This exterior entry door may be controlled by a key lock, card key, or
proximity reader. Entry into the animal room is via a double-door entry
which includes a change room and showerss. An additional double-door
access (air-lock) or double-doored autoclave may be provided for
movement of supplies and wastes into and out of the facility, respectively.
Doors to animal rooms open inward and are self-closing. Doors to
cubicles inside an animal room may open outward or slide horizontally or
vertically.
3. The animal facility is designed, constructed, and maintained to facilitate
cleaning and housekeeping. The interior surfaces (walls, floors, and
ceilings) are water resistant. Penetrations in floors, walls and ceiling
surfaces are sealed and openings around ducts and the spaces between
doors and frames are capable of being sealed to facilitate decontamination.
4. A hands-free or automatically operated hand washing sink is provided in
each animal room near the exit door. The sink trap is filled with an
appropriate disinfectant after each use.
5. Internal facility appurtenances, such as light fixtures, air ducts, and utility
pipes, are arranged to minimize horizontal surface areas.
6. Windows are not recommended. Any windows must be resistant to
breakage and must be sealed.
7. If floor drains are provided, they are always filled with an appropriate
disinfectant.
8. Ventilation should be provided in accordance with criteria from the Guide
for Care and Use of Laboratory Animals, latest edition. A ducted exhaust
air ventilation system is provided. This system creates directional airflow
which draws air into the laboratory from "clean" areas and toward
"contaminated" areas. The exhaust air is not recirculated to any other area
of the building. Filtration and other treatments of the exhaust air may not
be required, but should be considered based on site requirements, and
specific agent manipulations and use conditions. The exhaust must be
dispersed away from occupied areas and air intakes, or the exhaust must
be HEPA-filtered. Personnel must verify that the direction of the airflow
(into the animal areas) is proper. It is recommended that a visual
monitoring device that indicates and confirms directional inward airflow
be provided at the animal room entry. Consideration should be given to
installing an HVAC control system to prevent sustained positive


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pressurization of the animal spaces. Audible alarms should be considered
to notify personnel of HVAC system failure.
9. HEPA-filtered exhaust air from a Class II biological safety cabinet can be
recirculated into the animal room if the cabinet is tested and certified at
least annually. When exhaust air from Class II safety cabinets is to be
discharged to the outside through the building exhaust air system, the
cabinets must be connected in a manner that avoids any interference with
the air balance of the cabinets or the building exhaust system (e.g., an air
gap between the cabinet exhaust and the exhaust duct). When Class III
biological safety cabinets are used, they should be directly connected to
the exhaust system. If the Class III cabinets are connected to the supply
system, it is done in a manner that prevents positive cabinet
pressurization.
10. Cages are washed in a cage washer. The mechanical cage washer has a
final rinse temperature of at least 1800F.
11. An autoclave is available which is convenient to the animal rooms where
the biohazard is contained. The autoclave is utilized to decontaminate
infectious waste before moving it to other areas of the facility.
12. If vacuum service (i.e., central or local) is provided, each service
connection should be fitted with liquid disinfectant traps and an in-line
HEPA filter, placed as near as practicable to each use point or service
cock. Filters are installed to permit in-place decontamination and
replacement.
13. Illumination is adequate for all activities, avoiding reflections and glare
that could impede vision.
14. The completed Biosafety Level 3 facility design and operational
procedures must be documented. The facility must be tested for
verification that the design and operational parameters have been met
prior to operation. Facilities should be re-verified at least annually against
these procedures as modified by operational experience.
15. Additional environmental protection (e.g., personnel showers, HEPA
filtration of exhaust air, containment of other piped services, and the
provision of effluent decontamination) should be considered if
recommended by the agent summary statement, as determined by risk
assessment of the site conditions, or other applicable federal, state, or local
regulations.


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Table 1: Summary of Recommended Biosafety Levels for Infectious Agents


BSL Agents Practices Safety Equipment Facilities
(Primary Barriers) (Secondary Barriers)
Not known to consistently Standard Microbiological Practices None required Open bench top
cause disease in healthy Sink for hand washing
adults
2 Associated with human BSL-1 practice plus: Primary barriers = Class I or II BSL-1 plus:
disease, hazard = Limited access BCSs or other physical Autoclave available
percutaneous injury, Biohazard warning signs containment devices used for all Single pass air with no
ingestion, mucous membrane "Sharps" precautions manipulations of agents that cause recirculation
exposure Biosafety manual defining splashes or aerosols of infectious
any needed waste materials; PPEs; laboratory coats;
decontamination or medical gloves; face protection as needed
surveillance policies
3 Indigenous or exotic agents BSL-2 practice plus: Primary barriers = Class I or II BSL-2 plus:
with potential for aerosol Controlled access BCSs or other physical Physical separation
transmission; disease may Decontamination of all containment devices for all open from access corridors
have serious or lethal waste manipulations of agents; PPEs; Self-closing, double-
consequences Decontamination of lab protective lab clothing; gloves; door access
clothing before laundering respiratory protection as needed Exhausted air not
Baseline serum recirculated
Decontamination of all Negative airflow into
effluent laboratory (single
pass)
No floor drains
4 Dangerous/exotic agents BSL-3 practice plus: Primary barriers = All procedures BSL-3 plus:
which pose high risk of life- Clothing change before conducted in Class III BCSs or Separate building or
threatening disease, aerosol- entering Class I or II BCSs in combination isolated zone
transmitted lab infections; or Shower on exit with full-body air-supplied, Dedicated supply and
related agents with unknown All material decontaminated positive pressure personnel suit exhaust, vacuum, and
risk of transmission on exit from facility decontamination
systems
Other requirements
outlined in the text


November, 2008 Biosafety Manual










Table 2: Summary of Recommended Biosafety Levels for Activities in Which Experimentally or Naturally Infected Vertebrate
Animals Are Used


BSL Agents Practices Safety Equipment Facilities
(Primary Barriers) (Secondary Barriers)
S Not known to Standard animal care and management As required for normal care of each Standard animal facility
consistently cause practices, including appropriate medical species No recirculation of exhaust air
disease in healthy surveillance programs Directional air flow recommended
adults Hand washing Sink recommended
2 Associated with ABSL-1 practice plus: ABSL-1 equipment plus barriers: ABSL-1 facility plus:
human disease, hazard Limited access containment equipment appropriate for Autoclave available
= percutaneous injury, Biohazard warning signs animal species; PPES; laboratory coats, Hand washing sink available in
ingestion, mucous "Sharps" precautions gloves, face and respiratory protection animal room
membrane exposure Decontamination of all as needed. Mechanical cage washer used
infectious wastes and of animal
cages prior to washing

3 Indigenous or exotic ABSL-2 practice plus: ABSL-2 equipment plus: ABSL-2 facility plus:
agents with potential Controlled access Containment equipment for Physical separation from access
for aerosol Decontamination of all waste housing animals and cage corridors
transmission; disease Cages decontaminated before dumping activities Self-closing, double-door access
may have serious or bedding removed Class I or II BCSs available Sealed penetrations
lethal consequences Disinfectant foot bath as for manipulative Sealed windows
needed procedures(inoculation, Autoclave available in facility
Decontamination of all effluent necropsy) that may create
infectious aerosols. PPEs:
appropriate respiratory
protection
4 Dangerous/exotic ABSL-3 practice plus: ABSL-3 equipment plus maximum ABSL-3 facility plus:
agents which pose Entrance through change room containment equipment (i.e. Class III Separate building or isolated zone
high risk of life- where personal clothing is BCS or partial containment equipment Dedicated air supply and exhaust,
threatening disease, removed and laboratory in combination with full body, air- vacuum and decontamination
aerosol-transmitted clothing is put on; shower on supplied positive-pressure personnel systems
lab infections; or exit suit) used for all procedures and Other requirements outlined in the
related agents with All wastes are decontaminated activities text
unknown risk of before removal from facility
transmission


November, 2008 29


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Agents List


The following agents have been listed according to the most appropriate Biological
Safety Level to be used. The list presented below is based upon the risk groups given in
Appendix B of the March 1996 Guidelines for Research Involving Recombinant DNA
Molecules (NIH Guidelines), the agent summary statements in the CDC/NIH publication,
Biosafety in Microbiological and Biomedical Laboratories (BMBL), 4th edition (1999),
guidance from state and local regulatory agencies, and recommendations of the CDC.

Please note that Biological Safety Levels are not inherent to an agent but are performance
recommendations and should be chosen after a risk assessment is completed.

A proper risk assessment takes into account the characteristics of the agent involved, the
activities to be performed, and the environment in which the work will be completed.
Therefore, certain agents may be used at different Biological Safety Levels depending
upon the circumstances. For instance, human clinical samples from HIV-positive
patients may be safely handled at BSL-2. Growth of HIV in culture should be performed
under BSL-3 containment. Biological Safety Levels may be higher or lower than what is
given below for a particular agent depending upon the circumstances of its use.

The Biological Safety Office (BSO) reviews all projects involving recombinant DNA,
infectious disease agents, and agents of concern to livestock and agriculture and will
assist you in the risk assessment process. Once the Institutional Biosafety Committee
(IBC) and/or the Biological Safety Office assigns a Biological Safety Level, it must be
adhered to unless new information to warrant a change, in most cases from peer-reviewed
literature, is provided. The IBC and/or BSO will review the literature and make an
adjustment, if warranted.

Biological Safety Level 1 (BSL-1)
Agents that are not associated with disease in healthy adult humans, are of minimal
potential hazard to laboratory personnel, and of minimal potential hazard to the
environment may be used at BSL-1. Agents that may be used at BSL-1 include
Lactobacillus spp., asporogenic Bacillus subtilis or Bacillus licheniformis, Escherichia
coli-K12 (cloning strains), Baculoviruses, and adeno-associated virus types 1 through 4
in low concentrations (<109 IP/ml)

Those agents not listed under Biological Safety Levels 2, 3 and 4 are not automatically or
implicitly classified as BSL-1; a risk assessment must be conducted based on the known
and potential properties of the agents and their relationship to agents that are listed.

Biological Safety Level 2 (BSL-2)
Agents to be used at BSL-2 are associated with human disease which is rarely serious
and for which preventive or therapeutic interventions are often available. They are of
moderate potential hazard to laboratory personnel and/or the environment.


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BSL-2 Bacterial Agents Including Chlamydia
* Acinetobacter baumannii (formerly Acinetobacter calcoaceticus)
* Actinobacillus
* Actinomyces pyogenes (formerly Corynebacterium pyogenes)
* Aeromonas hydrophila
* Amycolata autotrophica
* Archanobacterium haemolyticum (formerly Corynebacterium haemolyticum)
* Arizona hinshawii all serotypes
* Bacillus acnhu U, ii
* Bartonella henselae, B. quintana, B. vinsonii
* Bordetella including B. pertussis
* Borrelia recurrentis, B. burgdorferi
* Burkholderia (formerly Pseudomonas species) except those listed under BSL-3
* Campylobacter coli, C. fetus, C. jejuni
* Chlamydiapsittaci, C. trachomatis, C. pneumoniae
* Clostridium botulinum, Cl. chauvoei, Cl. haemolyticum, Cl. histolyticum, Cl. novyi,
Cl. septicum, Cl. tetani
* Corynebacterium diphtheriae, C. pseudotuberculosis, C. renale
* Dermatophilus congolensis
* Edwardsiella tarda
* El) iipehili i\ rhusiopathiae
* Escherichia coli all enteropathogenic, enterotoxigenic, enteroinvasive and strains
bearing K1 antigen, including E. coli 0157:H7
* Haemophilus ducreyi, H. influenzae
* Helicobacterpylori
* Klebsiella all species except K. oxytoca (BSL-1)
* Legionella including L. pneumophila
* Leptospira interrogans all serotypes
* Listeria
* Moraxella
* Mycobacterium (except those listed under BSL-3) including M. avium complex, M.
asiaticum, M. bovis BCG vaccine strain, M. chelonei, M. fortuitum, M. kansasii, M.
leprae, M. malmoense, M. marinum, M. paratuberculosis, M. scrofulaceum, M.
simiae, M. szulgai, M. ulcerans, M. xenopi
* Mycoplasma, exceptM. mycoides andM. agalactiae which are restricted animal
pathogens
* Neisseria gonorrhoeae, N. meningitidis
* Nocardia asteroides, N. brasiliensis, N. otitidiscaviarum, N. transvalensis
* Rhodococcus equi
* Salmonella including S. arizonae, S. cholerasuis, S. enteritidis, S. gallinarum-
pullorum, S. meleagridis, S. paratyphi, A, B, C, S. typhi, S. typhimurium
* .1/ngel// including S. boydii, S. dysenteriae, type 1, S. flexneri, S. sonnei
* Sphaerophorus necrophorus
* Staphylococcus aureus


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* Streptobacillus moniliformis
* Streptococcus including S. pneumoniae, S. pyogenes
* Treponemapallidum, T. carateum
* Vibrio cholerae, V. parahemolyticus, V. vulnificus
* Yersinia enterocolitica
BSL-2 Fungal Agents
* Blastomyces dermatitidis
* Cladosporium bantianum, C. (Xylohypha) trichoides
* Cryptococcus neoformans
* Dactylaria galopava (Ochroconis gallopavum)
* Epidermophyton
* Exophiala (Wangiella) dermatitidis
* Fonsecaea pedrosoi
* Microsporum
* Paracoccidioides braziliensis
* Penicillium marneffei
* Sporothrix schenckii
* Trichophyton
BSL-2 Parasitic Agents
* Ancylostoma human hookworms including A. duodenal, A. ceylanicum
* Ascaris including Ascaris lumbricoides suum
* Babesia including B. divergens, B. microti
* Brugia filaria worms including B. malayi, B. timori
* Coccidia
* Cryptosporidium including C. parvum
* Cysticercus cellulosae (hydatid cyst, larva of T. solium)
* Echinococcus including E. granulosis, E. multilocularis, E. vogeli
* Entamoeba histolytica
* Enterobius
* Fasciola including F. gigantica, F. hepatica
* Giardia including G. lamblia
* Heterophyes
* Hymenolepis including H. diminuta, H. nana
* Isospora
* Leishmania including L. braziliensis, L. donovani, L. ethiopia, L. major, L. mexicana,
L. peruvania, L. tropica
* Loa loa filaria worms
* Microsporidium
* Naegleriafowleri
* Necator human hookworms including N. americanus
* Onchoerca filaria worms including, 0. volvulus
* Plasmodium including simian species, P. cynomologi, P. falciparum, P. malariae, P.
ovale, P. vivax
* Sarcocystis including S. sui hominis


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* Schistosoma including S. haematobium, S. intercalatum, S. japonicum, S. mansoni, S.
mekongi
* Sin. .i -hi, li e including S. stercoralis
* Taenia solium
* Toxocara including T canis
* Toxoplasma including T gondii
* Trichinella spiralis
* Trypanosoma including T brucei brucei, T brucei gambiense, T brucei rhodesiense,
T cruzi
* Wuchereria bancrofti filaria worms
BSL-2 Viruses
Adenoviruses, human all types

Alphaviruses (Togaviruses) Group A Arboviruses
* Eastern equine encephalomyelitis virus
* Venezuelan equine encephalomyelitis vaccine strain TC-83
* Western equine encephalomyelitis virus

Arenaviruses
* Lymphocytic choriomeningitis virus (non-neurotropic strains)
* Tacaribe virus complex
* Other viruses as listed in the BMBL

Bunyaviruses
Bunyamwera virus
Rift Valley fever virus vaccine strain MP-12
Other viruses as listed in the BMBL

Calciviruses

Coronaviruses

Flaviviruses (Togaviruses) Group B Arboviruses
Dengue virus serotypes 1, 2, 3, and 4
Yellow fever virus vaccine strain 17D
Other viruses as listed in the BMBL

Hepatitis A, B, C, D, and E viruses

Herpesviruses except Herpesvirus simiae (Monkey B virus), BSL-4
Cytomegalovirus
Epstein Barr virus
Herpesvirus ateles
Herpesvirus saimiri
Herpes simplex types 1 and 2
Herpes zoster


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Human herpesvirus types 6 and 7
Marek's disease virus
Murine cytomegalovirus
Pseudorabies virus

Orthomyxoviruses
Influenza viruses types A, B, and C
Other tick-borne orthomyxoviruses as listed in the BMBL

Papovaviruses
* All human papilloma viruses
* Bovine papilloma virus
* Polyoma virus
* Shope papilloma virus
* Simian virus 40 (SV40)

Paramyxoviruses
Newcastle disease virus
Measles virus
Mumps virus
Parainfluenza viruses types 1, 2, 3, and 4
Respiratory syncytial virus

Parvoviruses
Human parvovirus (B19)

Picornaviruses
Coxsackie viruses types A and B
Echoviruses all types
Polioviruses all types, wild and attenuated
Rhinoviruses all types

Poxviruses
Vaccinia all types except Monkeypox virus (BSL-3) and restricted poxviruses including
Alastrim, Smallpox, and Whitepox (restricted to the CDC, Atlanta, GA)

Reoviruses all types including Coltivirus, human Rotavirus, and Orbivirus (Colorado
tick fever virus)

Retroviruses
Avian leukosis virus
Avian sarcoma virus
Bovine leukemia virus
Clinical samples from HIV-positive patients
Feline immunodeficiency virus
Feline leukemia virus
Feline sarcoma virus


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Gibbon leukemia virus
Mason-Pfizer monkey virus
Mouse mammary tumor virus
Murine leukemia virus
Murine sarcoma virus
Rat leukemia virus

NOTE: Murine Retroviral Vectors
Murine retroviral vectors to be used for human transfer experiments (less than 10 liters)
that contain less than 50% of their respective parental viral genome and that have been
demonstrated to be free of detectable replication competent retrovirus can be maintained,
handled, and administered, under BL1 containment.

Rhabdoviruses
Rabies virus all strains
Vesicular stomatitis virus laboratory adapted strains ONLY including VSV-Indiana,
San Juan, and Glasgow
Togaviruses (see Alphaviruses and Flaviviruses)
Rubivirus (rubella)


Biological Safety Level 3 (BSL-3)
Agents to be used at BSL-3 are associated with serious or lethal human disease for which
preventive or therapeutic interventions may be available.
BSL-3 Bacterial Agents Including Rickettsia
* Bartonella
* Brucella including B. abortus, B. canis, B. suis
* Burkholderia (Pseudomonas) mallei, B. pseudomallei
* Coxiella burnetii
* Francisella tularensis
* Mycobacterium bovis (except BCG strain, BSL-2), M. tuberculosis
* Pasteurella multocida type B -"buffalo" and other virulent strains
* Rickettsia akari, R. australis, R. canada, R. conorii, R. prowazekii, R. rickettsii, R,
siberica, R. tsutsugamushi, R. typhi (R. mooseri)
* Yersinia pestis
BSL-3 Fungal Agents
* Coccidioides immitis (sporulating cultures; contaminated soil)
* Histoplasma capsulatum, H. capsulatum var.. duboisii
BSL-3 Parasitic Agents
None
BSL-3 Viruses and Prions
Alphaviruses (Togaviruses) Group A Arboviruses
* Semliki Forest virus
* St. Louis encephalitis virus


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* Venezuelan equine encephalomyelitis virus (except the vaccine strain TC-83 is BSL-
2)
* Other viruses as listed in the BMBL

Arenaviruses
* Lymphocytic choriomeningitis virus (LCM) (neurotropic strains)
* Flexal

Bunyaviruses
* Hantaviruses including Hantaan virus
* Rift Valley fever virus

Flaviviruses (Togaviruses) Group B Arboviruses
* Japanese encephalitis virus
* Yellow fever virus
* Other viruses as listed in the BMBL

Poxviruses
* Monkeypox virus

Prions
* Transmissible spongiform encephalopathies (TME) agents, Creutzfeldt-Jacob disease
and kuru agents (see BMBL for specific containment instruction)

Retroviruses
* Human immunodeficiency virus (HIV) types 1 and 2
* Human T cell lymphotropic virus (HTLV) types 1 and 2
* Simian immunodeficiency virus (SIV)

Rhabdoviruses
* Vesicular stomatitis virus


Biological Safety Level 4 (BSL-4)
Agents to be used at BSL-4 are likely to cause serious or lethal human disease for which
preventive or therapeutic interventions are not usually available.
BSL-4 Bacterial Agents
None
BSL-4 Fungal Agents
None
BSL-4 Parasitic Agents
None
BSL-4 Viral Agents
Arenaviruses (Togaviruses) Group A Arboviruses
* Guanarito virus


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* Lassa virus
* Junin virus
* Machupo virus
* Sabia virus

Bunyaviruses (Nairovirus)
* Crimean-Congo hemorrhagic fever virus

Filoviruses
* Ebola virus
* Marburg virus

Flaviruses (Togaviruses) Group B Arboviruses
* Tick-borne encephalitis virus complex including Absetterov, Central European
encephalitis, Hanzalova, Hypr, Kumlinge, Kyasanur Forest disease, Omsk
hemorrhagic fever, and Russian spring-summer encephalitis viruses

Herpesviruses (alpha)
* Herpesvirus simiae (Herpes B or Monkey B virus)

Paramyxiviruses
* Equine morbillivirus

Hemorrhagic fever agents and viruses as yet undefined.


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2 -- Information for Researchers








Project Registration
Some research projects involve work with potentially hazardous biological agents,
known infectious disease agents, or biological materials regulated by the federal or state
government. Many granting agencies require that the university monitor the use of
biological hazards, infectious disease agents, and recombinant DNA in order for them to
release funds to investigators. Therefore, we have developed a registration system to
ensure that all biological materials are handled properly and disposed of appropriately.
The Biological Safety Office administers four registration programs for research projects.

Bio-Agent (BA) Registration
Use of the following materials requires that the principal investigator completes and
submits the bio-agent registration document for approval by the Biological Safety Office.

Agents to be used at Biosafety Level 2 (BSL-2) or Biosafety Level 3 (BSL-3):
1. All human, animal, or plant pathogens that require BSL-2 or BSL-3 containment
and handling (see previous section: "Agents List") must be registered. Please
note that BSL-4 agents may not be used at UF.
2. Unknown human and animal pathogens must be registered. These are considered
BSL-2 until identified.
3. Cell lines or cultures that
1) have been immortalized with a virus (such as EBV or a retrovirus),
2) are known to be tumorigenic in primates (including humans), or
3) are primary human tumor cells.
These are considered BSL-2 (or higher in many cases).
4. Human blood or other tissues that are known to be HIV positive (or positive for
any human disease-causing virus or other agent), when used in research, must be
registered.

Recombinant DNA (R-DNA) Registration
All R-DNA projects that involve a living recombinant organism (this excludes projects
that involve DNA only, i.e. PCR) require registration with the Biological Safety Office.
A subset of R-DNA projects requires review and approval from the Institutional
Biosafety Committee (IBC). The UF IBC oversees all research projects and issues
involving R-DNA at UF. Use of the following requires that the principal investigator
completes and submits an R-DNA registration document.
1. All R-DNA projects, including the growth of recombinant bacteria for probe
isolation plasmidd or phage preparations) require registration. Projects must be
registered regardless of where the material came from or who originally
constructed it.
2. Projects that are exempt from the NIH Guidelines must also be registered.
3. The development of transgenic animals and plants requires registration.

R-DNA projects are performed at BSL-1, BSL-2, BSL-3 or the corresponding levels for
whole plant (BSL-1P, BSL-2P, BSL-3P) or whole animal (BSL-1N, BSL-2N, BSL-3N)
work. The Biological Safety Office, in conjunction with the IBC, will make the final
determination.


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Acute Toxins (AT) Registration
The use and storage of chemicals with a mammalian LD50 of < 100 [tg/kg. For a partial
list, see the Toxins Table that follows.

Regulated Biological Materials
Agents, such as plant pathogens or exotic microorganisms, that are regulated by federal
or state agencies (USDA/APHIS, EPA, FDA, DPI, etc.) shall be registered with the
Biological Safety Office by submission of a biological agent registration form and a
photocopy of the permit and permit conditions that have been granted by that agency. No
special form is required unless the agent fits into one of the first three categories.


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Table 3: Toxin Table


Toxins with a mammalian LD50 of 100 tLg/kg must be registered with the Biological
Safety Office. Therefore, use of the following toxins may require registration. If a toxin
is not on the list, it still may require registration, depending upon the LD50. For more
information, please contact the Biological Safety Office at 392-1591.

Toxicity

LD5o (ltg/kg)*

Abrin 0.7
Aerolysin 7.0
Botulinin toxin A 0.0012
Botulinin toxin B 0.0012
Botulinin toxin Cl 0.0011
Botulinin toxin C2 0.0012
Botulinin toxin D 0.0004
Botulinin toxin E 0.0011
Botulinin toxin F 0.0025
P-bungarotoxin 14.0
Caeruleotoxin 53
Cereolysin 40-80
Cholera toxin 250
Clostridium difficile enterotoxin A 0.5
Clostridium difficile cytotoxin B 220
Clostridium perfringens lecithinase 3
Clostridium perfringens kappa toxin 1500
Clostridium perfringens perfringolysin O 13-16
Clostridium perfringens enterotoxin 81
Clostridium perfringens beta toxin 400
Clostridium perfringens delta toxin 5
Clostridium perfringens epsilon toxin 0.1
Conotoxin 12-30
Crotoxin 82
Diphtheria toxin 0.1
Listeriolysin 3-12
Leucocidin 50
Modeccin 1-10
Nematocyst toxins 33-70
Notexin 25
Pertussis toxin 15


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Toxicity

LD5o (ltg/kg)*

Pneumolysin 1.5
Pseudomonas aeruginosa toxin A 3
Ricin 2.7
Saxitoxin 8
Shiga toxin 0.250
.\1/gel// dysenteriae neurotoxin 1.3
Streptolysin O 8
Staphylococcus enterotoxin B 25
Staphylococcus enterotoxin F 2-10
Streptolysin S 25
Taipoxin 2
Tetanus toxin 0.001
Tetrodotoxin 8
Viscumin 2.4-80
Volkensin 1.4
Yersinia pestis marine toxin 10

*Please note that the LD50 values are from a number of sources (see below). For
specifics on route of application (i.v., i.p., s.c.), animal used, and variations on the listed
toxins, please go to the references listed below.

Reference:
1. Gill, D. Michael; 1982; Bacterial toxins: a table of lethal amounts;
Microbiological Reviews; 46: 86-94
2. Stirpe, F.; Luigi Barbieri; Maria Giulia Battelli, Marco Soria and Douglas A.
Lappi; 1992; Ribosome-inactivating proteins from plants: present status and
future prospects; Biotechnology; 10: 405-412
3. Registry of toxic effects of chemical substances (RTECS): comprehensive guide
to the RTECS. 1997. Doris V. Sweet, ed., U.S. Dept of Health and Human
Services, Public Health Service, Centers for Disease Control and Prevention,
National Institute for Occupational Safety and Health; Cincinnati, Ohio


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Select Agents


The following lists of agents, toxins, and pathogens are classified by the Federal
government as Select Agents. Any possession, use, transfer or shipment of these
materials is strictly controlled by regulation.

See the EH&S web site at www.ehs.ufl.edu/bio for additional information and the UF
Select Agent Policy currently in place.

Researchers considering work with any of these materials must first contact the UF
Responsible Official at 392-1591 for the approvals, permits, clearances and other
necessary paperwork. Be aware that government clearance can take as much as 6 months
to complete in advance of any project.

Failure to comply with these Federal Regulations is punishable by both fines and
imprisonment.


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Select Agents List


HHS AND USDA SELECT AGENTS AND TOXINS
7 CFR Part 331, 9 CFR Part 121, and 42 CFR Part 73


HHS SELECT AGENTS AND TOXINS
Abrin
Cercopithecine herpesvirus 1 (Herpes B virus)
Coccidioides posadasii
Conotoxins
Crimean-Congo haemorrhagic fever virus
Diacetoxyscirpenol
Ebola virus
Lassa fever virus
Marburg virus
Monkeypox virus
Reconstructed replication competent forms of the 1918 pandemic
influenza virus containing any portion of the coding regions of
all eight gene segments (Reconstructed 1918 Influenza virus)
Ricin
Rickettsia prowazekii
Rickettsia rickettsii
Saxitoxin
Shiga-like ribosome inactivating proteins
South American Haemorrhagic Fever viruses
Flexal
Guanarito
Junin
Machupo
Sabia
Tetrodotoxin
Tick-borne encephalitis complex (flavi) viruses
Central European Tick-borne encephalitis
Far Eastern Tick-borne encephalitis
Kyasanur Forest disease
Omsk Hemorrhagic Fever
Russian Spring and Summer encephalitis
Variola major virus (Smallpox virus) and
Variola minor virus (Alastrim)
Yersinia pestis

OVERLAP SELECT AGENTS AND TOXINS
Bacillus anthracis
Botulinum neurotoxins
Botulinum neurotoxin producing species of Clostridium
Brucella abortus
Brucella melitensis
Brucella suis
Burkholderia mallei (formerly Pseudomonas mallei)
Burkholderia pseudomallei (formerly Pseudomonas pseudomallei)
Clostridium perfringens epsilon toxin
Coccidioides immitis
Coxiella bumetii
Eastern Equine Encephalitis virus
Francisella tularensis
Hendra virus
Nipah virus
Rift Valley fever virus
Shigatoxin
Staphylococcal enterotoxins
T-2 toxin
Venezuelan Equine Encephalitis virus


USDA SELECT AGENTS AND TOXINS
African horse sickness virus
African swine fever virus
Akabane virus
Avian influenza virus (highly pathogenic)
Bluetongue virus (Exotic)
Bovine spongiform encephalopathy agent
Camel pox virus
Classical swine fever virus
Cowdria ruminantium (Heartwater)
Foot-and-mouth disease virus
Goat pox virus
Japanese encephalitis virus
Lumpy skin disease virus
Malignant catarrhal fever virus
(Alcelaphine herpesvirus type 1)
Menangle virus
Mycoplasma capricolumi M.F38/M. mycoides Capri
(contagious caprine pleuropneumonia)
Mycoplasma mycoides mycoides
(contagious bovine pleuropneumonia)
Newcastle disease virus (velogenic)
Peste des petits ruminants virus
Rinderpest virus
Sheep pox virus
Swine vesicular disease virus
Vesicular stomatitis virus (Exotic)

USDA PLANT PROTECTION AND QUARANTINE (PPQ)
SELECT AGENTS AND TOXINS
Candidates Liberobacter africanus
Candidates Liberobacter asiaticus
Peronosclerospora philippinensis
Ralstonia solanacearum race 3, biovar 2
Schlerophthora rayssiae var zeae
Synchytrium endobioticum
Xanthomonas oryzae pv. oryzicola
Xylella fastidiosa (citrus variegated chlorosis strain)


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Maximum Toxin Amounts Excluded from Regulation

HHS Toxins Amount
Abrin 100mg
Conotoxin 100mg
Diacetoxyscirpenol (DAS) 1000mg
Ricin 100mg
Saxitoxin 100mg
Shiga-like ribosome inactivating proteins 100mg
Tetrodotoxin 100mg



Overlap toxins Amount
Botulinum neurotoxins 0.5mg
Staphylococcal enterotoxins 5.0mg
Clostridium perfringens epsilon toxin 100mg
Shigatoxin 100mg
T-2 toxin 1000mg


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Minors in Research Laboratories or Animal Facilities


Unless enrolled as a University of Florida student, minors are not allowed to work or conduct
research in University of Florida research laboratories, greenhouses or animal facilities
except as identified specifically below. In addition, minors are prohibited from operating
farm machinery or state vehicles and from working in machine shops.

1. All Minors are prohibited from working or conducting research in the following:

a. Any laboratory or facility designated as BSL-3, ABSL-3 or higher for
recombinant or infectious organisms.
b. Any laboratory where select agents or explosives are used or stored.
c. Any Animal Care Services (ACS) housing or procedure area/lab/facility. See
ACS Policy # ACS-PY-012. (Note that this does not apply to individual
Principal Investigator animal research laboratories).

2. Minors are prohibited from working with any of the following materials.

a. Radioactive materials or radiation (X-rays)
b. Acute Toxins

3. Minors are allowed to work or conduct research in laboratories (not listed in #1 above) if
the following requirements are met in full:

a. The University of Florida EH&S Policy titled; Minors In Research
Laboratories Or Animal Facilities (www.ehs.ufl.edu/bio/minors.htm) has
been read and understood.
b. A minor's research proposal registration form
(www.ehs.ufl.edu/bio/MinorReg.pdf) is submitted to and approved by the
University of Florida, Division of Environmental Health and Safety, or the
Institutional Biosafety Committee. Included in this form is The Potential
Hazards information sheet which requires a parental/guardian signature
indicating he/she has read of potential risks
c. Hazard specific safety training is completed by the Principal
Investigator/Sponsor with the minor as approved by EH&S.
d. Personal protective equipment, specific to the hazard, is provided to the minor
with instructions for use and disposal.
e. The minor is supervised at all times while in the laboratory and never left alone.
f Hours of work comply with Federal Regulation 29 CFR 570.35.
g. The laboratory is in full compliance with all applicable University of
Florida safety programs and regulations.


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Biological Waste Disposal Policy
This policy is intended to provide guidance and insure compliance with the NIH/CDC
guidelines, the State of Florida Administrative Code 64E-6, and restrictions of the
Alachua County Landfill.

Categories

1) Infectious/potentially infectious/R-DNA
a) human pathogens
b) animal pathogens
c) plant pathogens
d) recombinant DNA
e) human and primate blood, blood products and other body fluids
f) human and primate tissue
g) any material containing or contaminated with any of the above (test tubes,
needles*, syringes, tubing, culture dishes, flasks, etc.)

This waste must be inactivated prior to disposal. The preferred method is steam
sterilization (autoclaving), although chemical inactivation or incineration may be
appropriate in some cases. Storage of non-inactivated waste is restricted to within the
generating laboratory. The material may not be stored longer than 24 hours prior to
inactivation.
2) Non-infectious waste
This category includes waste that is not contaminated with any of the biological wastes
listed in category 1. It includes solid waste and sharps generated in clinical or laboratory
settings. Sterile or unopened biomedical materials that require disposal are also
considered biological waste.

IV packs test tubes petri dishes
needles* razor blades* tissue culture flasks
syringes culture dishes
scalpels* flasks
broken glass and plasticware** pipettes

This material does not require sterilization prior to disposal.
*must be packaged in plastic sharps boxes.
**must be within a box or other puncture proof container before adding to waste.
3) Mixed radioactive/biohazardous waste
The biohazardous component of mixed radioactive/biohazardous waste shall be
inactivated prior to its release to Radiation Safety for disposal as radioactive waste.
Steam-sterilization or chemical inactivation shall be employed as above. Although many
radioactive materials can be autoclaved safely, please check with the Radiation Safety


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Office (392-7359) regarding the best method to employ with any given radionuclide.
4) Mixed chemical/biohazardous waste
The biohazardous component of mixed chemical/biohazardous waste shall be inactivated
prior to its release for chemical disposal. Precautions should be taken to prevent the
generation and release of toxic chemicals during the inactivation process. In general,
autoclaving is not recommended because flammable or reactive compounds should not be
autoclaved due to the explosion hazard. Please check with the Biological Safety Office
(392-1591) for guidance regarding particular chemicals.
5) Animal carcasses and materials
The disposal of animal carcasses and other animal materials shall be
through the Animal Care Services incinerators or the Veterinary Medicine
tissue digesters only. These incinerators and digesters are for animal
materials only. Please contact Animal Care Services (392-2977) for
further information. No animal bodies or material shall be disposed of as
regular trash or through the biomedical waste receptacle.
6) Human remains
Please contact the State Anatomical Review Board (392-3588) for information regarding
the final disposition of human remains and body parts.

Packaging

1) Biohazard bags
These are used for the initial collection of certain biological wastes.

All biohazard bags must meet impact resistance (165 grams), tearing resistance (480
grams), and heavy metal concentration (<100 PPM total of lead, mercury, chromium and
cadmium) requirements. Written documentation (a test report) from the manufacturer
regarding these requirements must be on file. These bags must be placed in cardboard
boxes (see #3, below) prior to disposal.
2) Sharps
SNeedles, scalpels and razor blades are required to be containerized in
red plastic sharps containers. These are provided by Building Services
(392-4414) at the Health Center. All other sharps (broken glass and
plasticware, pipettes, etc.) shall be containerized in puncture-resistant
cardboard boxes (see #3, below). These are also available from
Building Services.
3) Corrugated cardboard boxes
All biological waste must be containerized in rigid, leak proof, puncture resistant boxes
as the terminal receptacle. The appropriate boxes are available from Building Services at
the Health Center, 392-4414.

Labeling
All packages containing biological waste shall be labeled with indelible ink marker (i.e.,
Sharpie) as follows:


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1) Date
Biohazard bags shall be labeled with the date they were put into use. Please note that
biohazard bags must be labeled even though they will be placed inside a secondary
container for final disposal.







Sharps containers shall be labeled with the date the container is full.
Corrugated boxes (biomedical waste boxes) shall be labeled with the date the
biohazardous waste was treated. Boxes used for non-biohazardous waste collection shall
be dated when the box is sealed.
2) Name/Location/Phone Number
Generator's (principal investigator's) name, lab location (room number) and phone
number will be clearly printed on each container.
3) Biohazard sign
Only manufactured containers with the preprinted universal biohazard symbol and the
words "biomedical," "biohazardous," or "infectious" shall be used.

Transport
The transport of biohazardous waste outside of the laboratory (i.e., to an autoclave or
incinerator) must be in a closed, leak-proof container that is labeled "biohazard." Only
trained personnel may transport biomedical waste. Labeling may be accomplished by use
of a red biohazard bag or a biomedical waste box with the universal biohazard symbol.
Only corrugated biomedical boxes and red plastic sharps containers may be used to
transport biological waste to the biomedical waste receptacle. Waste receptacle personnel
are instructed not to accept any other type of containers.

Transportation of red-bagged waste must be in closed, leak-proof containers, properly
labeled as "Biohazards." Movement of regulated/biological waste through public
corridors, along carpeted hallways, and on public elevators must be avoided. Any
leakage/spills from these containers must be immediately reported to the Biosafety Office
at 392-1591. Signs must be displayed to prevent tracking of the spills to other areas.

Training
All employees who handle biological waste shall be trained annually regarding its proper
handling. All new employees shall be trained before they are allowed to handle
biological waste.

Training may be accomplished through the UF Bloodborne Pathogen Training Program,
informally in the lab setting, or through formal training programs set up by individual
departments or divisions. For assistance, please call the Biological Safety Office at
(392-1591).

According to Florida Statute (Ch. 64E-16 F.A.C.), records of the training session shall be
maintained for each employee, along with an outline of the training program. Training
records shall be retained for a period of three (3) years.


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Biological Waste Disposal Containers
The following waste disposal containers for biohazardous or biomedical waste are to be
used for teaching and research purposes at the University of Florida:

ALL BIOHAZARDOUS WASTE MUST BE TREATED BEFORE DISPOSAL

Biohazard Boxes
Biohazard boxes lined with red bags shall be used to dispose of treated (autoclaved or
bleached) biomedical waste. These boxes are available from HSC Building Services
(392-4414). Boxes shall be labeled with principal investigator's name, phone number,
date of treatment, and lab location (room number). The 3A full boxes shall be labeled
with indelible ink markers (e.g. Sharpie). The 3 full boxes shall be sealed with a leak-
resistant, clear, strapping tape in an H pattern. All edges where leakage could occur shall
be taped. Trained research or custodial staff only shall handle the full boxes.

H-Pattern Taping Sequence










1st 2n 3rd


Disposal of biomedical waste in the regular trash or dumpster is prohibited. The Alachua
County Landfill does not accept any items that resemble "biomedical devices" and will
reject the entire load. In some cases, the individual principal investigator has been made
responsible for the costs incurred for sorting and disposal of improperly disposed waste.

Sharps Boxes
Red plastic sharps boxes are used for disposal of needles, razor blades,
scalpels, and small Pasteur pipettes. These are available from HSC
Building Services (2-4414). Sharps boxes that contain infectious
materials must be inactivated by autoclaving the closed box. These boxes
shall be labeled (with indelible ink marker as above) with the principal
investigator's name, phone number, date the sharps box is full, and lab
location (room number). Trained research or custodial staff only shall
handle the full sharps boxes.


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Biohazard Bags
Red plastic biohazard bags shall be used for collection of biohazardous tissue culture
items, petri dishes, and other non-sharp items. These biohazard bags shall be
immediately labeled with an indelible ink marker (e.g. Sharpie@) when the bag is first
put into use. Labels shall include the principal investigator's name, phone number, the
date the first biohazard item is placed into the bag, and lab location (room number). The
biohazardous waste items must be inactivated by autoclaving in biohazard autoclave bags
or treatment with bleach within 30 days of accumulation (Ch. 64E-16 F.A.C.). After
treatment, the bags shall be tightly sealed and placed in biohazard boxes. They may be
transported to the biomedical waste receptacle by trained laboratory personnel or by
trained custodial staff. Biohazard bags should never be handled by non-research staff or
placed in the hallway. Double bagging may be required to prevent leakage, however,
absorbent material must be added to prevent the presence of any liquid and fluid. Full,
untreated biohazard bags shall be stored only in the lab where accumulation occurred.
Full biohazard bags must be treated within 24 hours.










Fold box flaps down. Seal bag by twisting Double upper section
Insert labeled liner and fold and taping. over and tape to
over edges. double seal the bag.

Autoclave Use and Testing
Rules governing the use and testing of autoclaves are based on Rule 64E-16 of the
Florida Administrative Code.

Autoclave Testing
Autoclaves shall be tested before being placed into service and then periodically for
effectiveness.
1) Periodicity
a) Every 40 hours of use
Required for autoclaves that are used to inactivate human or non-human
primate blood, tissues, clinical samples, or human pathogens.
b) Every 6 months
Required for autoclaves that are used to inactivate other material.
2) Method
A commercially available test indicator kit that uses bacterial spores (Geobacillus
stearothermophilus) is the approved method of testing autoclave efficiency. Most
spore vial test kits require 56C to 600C incubation of the autoclaved test vial
Biosafety Manual November, 2008 51







along with a non-autoclaved control vial. Incubation causes surviving spores to
grow.

a) New autoclaves
Before placing an autoclave into service, a test load approximating the
weight and density of the type of waste generated shall be autoclaved with
test spore vials. The spore vials should be placed at the bottom, top,
front, rear, and center of the autoclave chamber. This can be achieved by
either:
-placing vials at those positions within one large test load, OR
-making several smaller test packs with 1 vial at the center of each
and placing the packs at those locations within the chamber.
The appropriate parameters for sterilization including temperature, pressure, and
treatment time shall be determined in this way.
b) Autoclaves already in use
For periodic testing, place a spore vial in the very center of a test load
prior to autoclaving.
3) Storage Information
Please read the product information sheet for appropriate storage information.
Spore vials should not be frozen. Each batch of vials has an expiration date.
Vials should not be used after their expiration date.


Record-Keeping
The following records regarding autoclave use must be kept:
1) On-site maintenance records
2) Autoclave use log must be available near the autoclave.
Each load of material inactivated shall be logged as follows:
a) Date, time, and operator's name
b) Contact Information: Lab, room number, phone number
c) Is this biohazardous material?
d) Confirmation of sterilization:
i) Record the temperature, pressure, and length of time the load is
sterilized. Please note that temperature sensitive autoclave tape is
not sufficient to indicate that the load reached sterilization
conditions because the tape will change color at lower
temperatures.
ii) Save the autoclave printout, if the autoclave has a working printer.


Autoclave Operating Procedures
A written sterilization procedure shall be in place for each workplace. This shall include
the following:
1) Parameters
Appropriate parameters for sterilization shall be determined from the testing with
spore vials. The time it takes to sterilize a load will change depending upon the
load density and the sterilization cycle one chooses. Tests have been performed
which imitate these various situations. Please follow the established guidelines.
2) Protocol


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Identification of standard treatment containers and proper load placement shall be
made.



3) Cleaning
The autoclave and work areas shall be cleaned after every use and the work area
shall be disinfected, as needed.


Autoclave Operation and Safety Training
Autoclave training is available from the EH&S Biological Safety Office both quarterly
and upon request. The training is geared toward the research staff and goes over proper
use of autoclaves and how they may be maintained and used properly. Safety training is
also given.


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Autoclave Guidelines


AUTOCLAVE GUIDELINES

STERILIZATION TIMES (Drying Time Not Included)

1210C / 2500F and 15 p.s.i.


BIO WASTE IN AUTOCLAVE BAGS, LOOSELY TIED

MULTIPLE BAGS --- 100 MINUTES OR LONGER

SINGLE BAG (FULL) --- 90 MINUTES

PARTIAL BAG --- 60 MINUTES



DRY GOODS


GLASSWARE, EMPTY, INVERTED

INSTRUMENTS, WRAPPED

UTENSILS, WRAPPED


--- 15 MINUTES

--- 30 MINUTES

--- 30 MINUTES


LIQUIDS (Bottles with vented caps 12 FULL)

75mL --- 25 MINUTES
250mL --- 30 MINUTES
500mL --- 40 MINUTES
1000mL --- 45 MINUTES
1500mL --- 50 MINUTES
2000mL --- 55 MINUTES


The above times should be used as a guide in determining the length of time items should
be autoclaved in order to achieve sterilization.

Questions? Please call Gary Smith at 392-1591


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Disinfectants


"Disinfectant" refers to an agent that is applied to treat (usually) inanimate objects to
render them free of pathogenic or infectious non-sporeforming microorganisms. In
contrast, the term "sterilant" refers to an agent that renders items free of all
microorganisms, including spores. The two are not the same and should not be confused.

Disinfectants are used in laboratory and chemical settings to 1) treat a surface or an item
before or after routine use, or 2) to treat a surface or an item after a spill or
"contaminating event."

Because disinfectants are antimicrobial, they may, by their nature, also have a toxic effect
to the user. Therefore, Material Safety Data Sheets and other manufacturer's product
information should be available and thoroughly reviewed before using these products.

There are many different types and formulations of disinfectants. The researcher or
clinician should ensure that the proper product, one that is effective against the specific
microorganism being studied, is used.

The FDA regulates those products that are marketed as sterilants or sanitizing agents on
medical devices. They have published a list of products currently on the market that are
labeled as sterilants (http://www.epa.gov/spdpublc/snap/sterilants/index.html).

The EPA regulates moderate and low level disinfectants as "chemical germicides" under
their pesticide regulations. They have published a list of 1) those products that are
effective against Mycobacterium spp. (tubercle bacilli), and 2) those products that are
effective against HIV-1 (human immunodeficiency virus).

Please contact the Biological Safety Office for information about any of these lists or for
a list of manufacturers. Be aware that most disinfectants assume pre-cleaning to remove
gross organic/protein prior to use.

Whenever a disinfectant or sterilant is used, proper safety precautions must be followed.
Appropriate clothing (gloves, safety goggles, aprons) must be worn. In addition, these
compounds must be used in well-ventilated areas.

Following is a discussion of general categories of disinfectants. Please note that there are
several different products and different formulations in each category.

Liquids

Alcohols
The most commonly used alcohols, ethanol and isopropanol, are most effective at
concentrations of 70% (v/v) in water. Both higher and lower concentrations are less
effective. Alcohols are active against vegetative bacteria, fungi, and lipid containing
viruses but not against spores. Their action on lipid-containing viruses is variable.
Alcohols are difficult to use as contact disinfectants because they evaporate rapidly and
do not penetrate organic matter well. When using alcohols, it is best to clean an object,
then submerge it in alcohol for the appropriate time. Alcohols are often used in concert


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with other disinfectants such as formaldehyde (but see toxicity warning below) or
chlorine (2000 ppm chlorine in alcohol). Alcohol is NOT a registered tuberculocidal or
HIV listed disinfectant.
Chlorine compounds
The most commonly used and generally effective disinfectant is sodium hypochlorite
(common household bleach). It is a strong oxidizing agent and therefore can be corrosive
to metal. A 1:50 dilution, supplying 1000 ppm available chlorine, of the common
household product (e.g. Clorox) is very effective as a general laboratory disinfectant and
a 1:10 dilution supplying 5000 ppm available chlorine is effective against spills involving
blood or other organic material. Please note that the presence of high concentrations of
protein can inactivate the action of chlorine products. Dilute hypochlorite solution must
be prepared daily to be maximally effective. There are more concentrated sodium
hypochlorite solutions available for industrial use, so please read the product information
carefully to determine the proper dilution.

Table 4: Dilutions of Household Bleach

Volume Volume Dilution Sodium Available
S Of Bleach of Water Ratio Hypochlorite % Chlorine
(mg/L)
Undiluted 0 1:1 5.25 50,000
1 9 1:10 0.5 5,000
1 99 1:100 0.05 500
Formaldehyde
Formaldehyde is a gas that is available either dissolved in water and methanol as a 37%
formaldehyde solution, known as formalin, or as a solid, paraformaldehyde, that may be
melted to release the gas. Formaldehyde gas kills all microorganisms and spores but not
prions. It is used for space decontamination and to decontaminate biological safety
cabinets, a dangerous process requiring specifically trained personnel. Formaldehyde
dissolved in water is active at 1-8% solutions and can be used to decontaminate hard
surfaces. However, because formaldehyde is an irritant at low concentrations (0.1 to 5
ppm) and a probable carcinogen, its use as a hard surface disinfectant is limited to
situations in which it is particularly needed. Due to its toxic effects, there are no EPA-
registered disinfectants that contain formaldehyde.
Glutaraldehyde
Glutaraldehyde is usually supplied as a 2% solution and requires activation by the
addition of an alkaline agent prior to use. The activated product may be kept for about
two weeks and should be discarded when turbid. Glutaraldehyde is active against all
microorganisms, but is toxic, an irritant, and mutagenic and should be used only when
necessary. Please follow the manufacturer's guidance when using glutaraldehyde-based
products because there are many different formulations that have been designed for
specific uses.
Hydrogen peroxide
Hydrogen peroxide is usually available as a 30% solution. It may be diluted 1:5 for use
as a disinfectant to decontaminate work surfaces of laboratory benches and biosafety
cabinets. It is active against a wide array of microorganisms. However, it is an oxidizing


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agent and should not be used on aluminum, copper, zinc, or brass. Hydrogen peroxide is
unstable at high temperatures and in light.
Iodine and lodophors
Iodine and iodophors, compounds in which the iodine is combined with a solubilizing or
carrier agent, are general, all-purpose disinfectants with an action similar to that of
chlorine products. The appropriate concentration for iodine-containing products is 75
ppm available iodine for disinfecting work surfaces. Concentrations may be much higher
for other purposes. Like chlorine compounds, the effectiveness of iodine compounds
may be diminished in the presence of protein/organic material. Iodophor compounds that
are used for antisepsis (germicide applied to tissue or skin) are not appropriate for use as
hard surface disinfectants and vice versa. Please read the product material for
appropriate dilutions and applications.
Phenol and phenolic compounds
Phenolic compounds are active at 0.2 to 3% concentrations against all forms of
vegetative microorganisms but not against spores. They have only limited effectiveness
against non-lipid viruses and, when properly formulated, show anti-mycobacterial
activity. There are many common disinfectants based on phenol and they should be used
according to the manufacturer's recommendations.
Quaternary ammonium compounds
Compounds in this class are active at concentrations of 0.1 2%. They are active against
vegetative bacteria, lipid viruses, but not against bacterial spores, non-lipid viruses, or
tubercle bacilli. These compounds should be used only when a low-level disinfectant is
required.

Gases

Ethylene Oxide
Inactivates micro-organisms, cellular disruption; used only as surface sterilizer
Temp 500C-60C, Humidity 30-40%
Conc. 400-800 mg/liter
Time 2 hrs at 600C or 24 hrs at 200C
Aeration required after cycle for 8 hrs
Biological indicators needed to confirm kill
Suspected carcinogen with explosive properties
Vapor Phase Hydrogen Peroxide
Temp 40C-600C,> temp results in > activity
Conc. 30%, less than 10 mg/liter
Non-toxic end products of water and oxygen
Limited to surfaces, no penetration
Corrosive to some materials
Degrades natural rubber and nylon
Chlorine Dioxide gas
Dilute chlorine gas and sodium chlorite, less than 25 mg/liter
Temp 250C-30C, pre-humidification required
Limited to surfaces, no penetration
Corrosive to some materials
Mucous membrane irritant
Ozone
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Oxidizing properties, inhibits bacterial growth, reacts with amino acids,
RNA/DNA
Temp 25C
Conc. 2-5 mg/liter
Systems convert oxygen to ozone
Limited penetration
Formaldehyde Gas (from heating paraformaldehyde)
Temp 200C-220C, humidity 60-85%
Conc. 0.3 gm/cu ft of volume
Time 6-8 hours
Toxic irritant and suspected carcinogen
Limited penetration, primarily surface action
Requires aeration and time for formaldehyde to off-gas, usually 8 hours


Irradiation

Ultraviolet, UV radiation
Virucidal action correlates with shorter wavelengths of the UV spectrum,
250-260 nm.
Mechanism of UV radiation injury attributed to absorption by and resultant
damage to nucleic acids
Due to low energy, the power of penetration is poor.
Dust and thin layers of proteins on surfaces reduce the virucidal activity
Ionizing radiation
Penetrates products and micro-organisms
Releases high energy, disrupts cells, DNA & RNA and results in kill
Damage done to cell membrane and other cellular structures
Cobalt 60, most common technique used for commercial sterilization of single-
use items, and Cesium 137
Electron Beam
Limited penetration
Accelerator generates high energy electrons
Microwaves
Thermal and non-thermal activity
Still under study for the most part


Disinfectants Bibliography
The following materials were consulted or used in this chapter:
Disinfection, Sterilization, and Preservation. Fourth edition. 1991. Seymour S. Block ed., Lea & Fibiger,
Philadelphia
Laboratory Biosafety Manual. Third edition. 2004. World Health Organization, Geneva.
Biological Safety: Principles and practices. Third edition. 2000. Diane O. Fleming et al. eds. ASM Press,
Washington DC
Manual of Clinical Microbiology. Eighth edition. 2003. Patrick Murray ed., ASM press, Washington DC
Prudent Practices in the Laboratory: Handling and disposal of chemicals. 1995 National Research
Council. National Academy Press, Washington


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Table 5: Summary and Comparison of Liquid Disinfectants (Page 1)


Commonly used disinfectants, recommended when appropriate.


Class Recommended Use How They Work Advantages 'Disadvantages Comments/Hazards Examples
70% Isopropyl Cleaning some Changes protein Fairly inexpensive <50% solution not very eFlammable
alcohol instruments structure of effective *Eye Irritant
solution Cleaning skin microorganism Not active when organic *Toxic
Presence of water matter present
assists with killing action Not active against certain
types of viruses
Evaporates quickly contact
time not sufficient for killing
Chlorine Spills of human body Free available chlorine Kill hardy viruses Corrode metals such as Follow spill procedure Bleach
compounds fluids combines with contents (e.g., hepatitis) stainless, aluminum and dilution instructions solutions
Bactericidal Good within microorganism Kill a wide range of Organics may reduce Make fresh solutions (sodium
Fungicidal Good reaction byproducts cause organisms activity before use hypochlorite)
Sporicidal Good at its death Inexpensive Increase in alkalinity Eye, skin, and Clorox
>1000 ppm Sodium Need 500 to 5000 ppm Penetrates well decreases bactericidal property respiratory irritant Cryosan
Hypochlorite Produce chemical Relatively quick Unpleasant taste and odor *Corrosive Purex
combination with cell microbial kill Tuberculocidal with *Toxic
substances May be used on food extended contact time
Depend upon release of prep surfaces
hypochlorous acid
Gluteraldehyde Bactericidal Good Coagulates cellular Non-staining, Not stable in solution (shelf *Eye, skin and Calgocide
Fungicidal Good proteins relatively non- life 14 days) respiratory irritant. 14
Tuberculocidal corrosive Has to be in alkaline *Sensitizer Cidex
slow acting Useable as a sterilant solution *Toxic Vespore
Virucidal Good on plastics, rubber, Inactivated by organic
Sporicidal Good lenses, stainless steel, material
and other items that
can't be autoclaved


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Table 5: Summary and Comparison of Liquid Disinfectants (Page 2)
Class Recommended How They Work Advantages Disadvantages Comments/Hazards Examples
Use
lodophors Disinfecting some Free iodine enters Kill broad range of May stain plastics or Dilution critical follow Bactergent
(iodine with semi-critical medical microorganism and organisms corrode metal directions! Hy-Sine
carrier) equipment binds with its cellular Highly reactive May stain Use only EPA- Ioprep
Bactericidal Very components Low tissue toxicity skin/laundry registered hard surface Providone-
Good Carrier helps penetrate Kill immediately rather than Stains most materials iodophor disinfectants iodine;
Fungicidal Excellent soil/fat by prolonged period of stasis Odor Don't confuse skin betadine
Virucidal Excellent Need 30 to 50 ppm Not affected by hard water Some organic and antiseptic iodophors for *
Probably by disorder May be used on food prep inorganic substances disinfectants Wescodyne
of protein synthesis due surfaces neutralize effect *Skin and eye irritant
to hindrance and/or Tuberculocidal with *Corrosive
blocking of hydrogen extended contact time *Toxic
bonding Sporicidal some
Phenolic Bactericidal Gross protoplasmic Nonspecific concerning Unpleasant odor *Skin and eye irritant Hil-Phene
Compounds Excellent poison bactericidal and fungicidal Some areas have *Sensitizer Lph
Fungicidal Excellent Disrupts cell walls action disposal restrictions *Corrosive Metar
Tuberculocidal Precipitates cell When boiling water would Effectiveness reduced *Toxic Vesphene
Excellent proteins cause rusting, the presence of by alkaline pH, natural
Virucidal Excellent Low concentrations phenolic substances produces soap, or organic
inactivate essential an anti-rusting effect material
enzyme systems Sporicidal NO
Quaternary Ordinary Affect proteins and cell Contain a detergent to help Do not eliminate Select from EPA list of Coverage
ammonium housekeeping membrane of loosen soil spores, TB bacteria, hospital disinfectants 258
compounds (e.g., floors, microorganism Rapid action some viruses *Skin and eye irritant End-Bac
(QUATS) furniture, walls) Release nitrogen and Colorless, odorless Effectiveness *Toxic Hi Tor
Bactericidal phosphorous from cells Non-toxic, less corrosive influenced by hard
Excellent Highly stable water
Fungicidal Good May be used on food prep Layer of soap
Virucidal Good (not surfaces interferes with action
as effective as phenols) _____


Barbara Fox Nellis (8-23-96)


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Table 6: Summary of Practical Disinfectants


Quaternary Ammonium Phenolic Chlorine Isopropyl
Compounds Compounds Compounds lodophor Ethyl Alcohol Alcohol Formaldehyde Glutaraldehyde
Inactivates
Vegetative Bacteria + + + + + + + +
Lipoviruses + +a + + +a + + +
Nonlipid Viruses + + + +
Bacterial Spores + + + +
Treatment Requirements
Use Dilution 0.1-2.0% 1.0-5.0% 500ppmb 25-1600ppmb 70-85% 70-85% 0.2-0.8% 2%
Contact Time-minutes
Lipovirus 10 10 10 10 10 10 10 10
Broad Spectrum NE NE 30 30 NE NE 30 30
Important Characteristics
Effective Shelf Life + + + + + + +
> 1 week
Corrosive + + +
Flammable + +
Explosion Potential
Inactivated by organic + -+ +
matter
Skin Irritant + + + + + +
Eye Irritant + + + + + + + +
Respiratory Irritant + + +
Toxicd + + + + + + + +
Applicability
Waste Liquids +
Dirty Glassware + + + + + + + +
Equipment, Surface + + + + + + + +
Decontamination
Proprietary Productse A-33, CDQ, End-Bac, Hll-Phene, Chloramine Hy-Sine, loprep, Sterac Cidex
H1-Tor, Mikro-Quat Metar, Mikro- T, Clorox, Mikroklene,
SBac, O-Syl Purex Wescodyne___
Source: Adapted from Laboratory Safety Monograph, A Supplement to the NIH Guidelines for Recombinant DNA Research. National Institutes of Health, Office of
Research Safety, National Cancer Institute, and the Special Committee of Safety and Health Experts, Bethesda, Maryland. January 1989: 104-105
+ Yes a Variable results depending on virus d By skin or mouth or both
No b Available Halogen e Space limitations preclude listing all products available. Individual listings (or omissions)
NE Not Effective c Protected from light and air do not imply endorsement (or rejection) of any product by the National Institutes of Health.


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Table 7: Reprocessing Methods for Equipment Used in the Health Care Setting

Sterilization: Destroys: All forms of microbial life including high numbers of bacterial spores.

Steam under pressure (autoclave), gas (ethylene oxide), dry heat or immersion in an approved chemical "sterilant" (e.g., US Environmental
Methods: Protection Agency-approved) for prolonged period of time, e.g., 6-10 hours or according to manufacturer's instructions. Note: Liquid chemical
"sterilants" should be used only on those instruments that are impossible to sterilize or disinfect with heat.

For those instruments or devices that penetrate skin or contact normally sterile areas of the body, e.g., scalpels, needles, etc. Disposable invasive
Use: equipment eliminates the need to reprocess these types of items. When indicated, however, arrangements should be made with a health-care facility
for reprocessing of reusable invasive instruments.

High-Level Destroys: All forms of microbial life except high numbers of bacterial spores.
Disinfection:
Metho: Hot water pasteurization (80-100C, 30 minutes) or exposure to an approved (e.g., US EPA-approved) "sterilant" chemical as above, except for a
short exposure time (10-45 minutes or as directed by the manufacturer).

Use: For reusable instruments or devices that come into contact with mucous membranes (e.g., laryngoscope blades, endotracheal tubes, etc.).

Intermediate- Destroys: Mycobacterium tuberculosis, vegetative bacteria, most viruses and most fungi, but does not kill bacterial spores.
Level
Disinfection: Approved (e.g. US EPA-approved) "hospital disinfectant" chemical germicides that have a label claim for tuberculocidal activity; commercially
Methods: available hard-surface germicides or solutions containing at least 500 ppm free available chlorine (a 1:10 dilution of common household bleach-
about % cup bleach per gallon of tap water).

For those surfaces that come into contact only with intact skin, e.g., stethoscopes, blood pressure cuffs, splints, etc., and have been visibly
Use: contaminated with blood or bloody body fluids. Surfaces must be pre-cleaned of visible material before the germicidal chemical is applied for
disinfection.

Low-Level Destroys: Most bacteria, some viruses, some fungi, but no Mycobacterium tuberculosis or bacterial spores.
Disinfection:
Methods: Approved (e.g., US EPA-approved) "hospital disinfectants" (no label claim for tuberculocidal activity).

Use: Use these excellent cleaning agents for routine housekeeping or removal of soiling in the absence of visible blood contamination.

Environmental Environmental surfaces that have become soiled should be cleaned and disinfected using any cleaner or disinfectant that is intended for
Disinfection: environmental use. Such surfaces include floors, woodwork, ambulance seats, countertops, etc.

Important: To assure effectiveness of any sterilization or disinfection process, equipment and instruments must first be thoroughly cleaned of all visible soil


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Shipment of Biological Materials
The following regulations apply to the packaging and shipment of biological materials:

U.S. Department of Transportation, 49 CFR Parts 171-180 and amendments
U.S. Public Health Service, 42 CFR Part 72, Interstate Shipment of Etiologic Agents
U.S. Department of Labor, Occupational Safety and Health Administration, 29 CFR Part
1910.1030, Bloodborne Pathogens
International Air Transport Association (IATA), Dangerous Goods Regulations
U.S. Postal Service, 39 CFR Part 111, Mailability of Etiologic Agents, Mailability ofSharps and
Other Medical Devices, and Publication 52, Acceptance ofHazardous, Restricted or
Perishable Matter
International Civil Aviation Organization, Technical Instructions for the Safe Transport of
Dangerous Goods by Air
United Nations, Recommendations of the Committee ofExperts on the Transportation of
Dangerous Goods

All North American airlines and FedEx, the largest shipper of infectious materials, use
the IATA regulation (also referred to as the Dangerous Goods Regulation or DGR) as
their standard. Meeting the conditions of this standard will ensure meeting the provisions
of the other US regulations.

Many biological materials fall into the category of "dangerous goods" for shipping
purposes. All individuals involved in the transport of dangerous goods or the preparation
of dangerous goods for transport must be trained to do so properly and safely. In
addition, we require safe transport of items within facilities and around campus. These
topics are covered in the "Shipping and Transport of Biological Materials" training given
by the Biosafety Office.

You must have a training certificate from UF EH&S to ship biological materials.
Call us or see our list of scheduled classes at http://www.ehs.ufl.edu/Bio/default.asp.

Training is valid for 2 years and is a federal requirement designed to protect yourself,
your co-workers, the public drivers, airline staff, crew, pilots, passengers, and package
recipients.
Training also ensures successful shipments; carriers or Federal regulators may open,
delay, or reject your shipment if it's not correct. Training also prevents penalties for
violations; civil penalties range from $250 to $27,500 per violation per day, and criminal
penalties for willful violations range up to $500K and 5 years in jail.

Classes cover the following:

1. Classifying the material Is it regulated? Is it forbidden for transport?
2. Identifying the material select the proper shipping name
3. Choosing the right packaging
4. Packaging it correctly









5. Marking & Labeling the shipment correctly
6. Supplying additional required documentation dangerous goods declaration
forms
7. Making shipping arrangements i.e. permits, customs documents for overseas
shipments
8. Transporting things safely around UF hand carrying & vehicle transport



Biological materials subject to shipping & transport regulations:


In the context of shipping regulations, "Dangerous Goods" are "Articles or substances
which are capable ofposing a risk to health, safety, property or the environment &
which are shown in the list of dangerous goods in the Regulations or which are
classified according to these Regulations." (49 CFR Parts 100-185 & IATA 1.0).

Biological materials that fall under this definition include:

Biological toxins
Infectious substances
Diagnostic specimens
Biomedical waste
Cultures
Genetically Modified Organisms

Other biological material that may be regulated via state or federal permits:

Plants,
Plant pests
Insects
Cell cultures
Live animals

Items that frequently accompany shipments of biological material are also regulated:

Dry ice
Environmental pollutants (e.g. formalin)
Alcohol
Fixative solutions



Transporting biological material within and around UF:


The following general guidelines apply:


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Double contain the items plastic leak-proof containers within sturdy outer
packaging. Include absorbent material within the containers as well as padding to
minimize movement of the containers) within the outer packaging. Wipe the
outer container with an appropriate disinfectant before removing it from the
laboratory and apply a biohazard sticker if applicable. Put your name and contact
information on the package. Individuals transporting biohazardous agents should
be knowledgeable about handling spills.

UF policy states that dangerous goods are not to be transported in your personal
vehicle. This is both a safety and liability issue. Use a state vehicle.


Shipping radioactive? Call 392-7359
Shipping chemicals? Call 392-8400



Permits:
Federal or state permits may be required for some biological materials. See below or
contact us at 392-1591 for more information. Permits are issued in the name of the PI
who is required to keep them updated and current as necessary; the Biosafety Office does
not hold any "centralized" permits.

1. CDC Import Permit http://www.cdc.gov/od/eaipp/ for import of etiological agents
causing disease in humans, non-sterilized human or animal tissues/fluids known
or suspected to contain disease agents, hosts/vectors known or suspected to
contain disease agents

2. USDA/APHIS Veterinary Permit http://www.aphis.usda.gov/vs/ncie/ needed for
import of materials derived from (livestock/poultry) animals or exposed to
(livestock/poultry) animal-source materials, including: animal tissues, blood, cells
or cell lines of livestock or poultry origin, RNA/DNA extracts, hormones,
enzymes, monoclonal antibodies for IN VIVO use in non-human species, certain
polyclonal antibodies and antisera, bulk shipments of test kit reagents, arthropod
vectors of livestock diseases, and microorganisms infectious to livestock
including bacteria, viruses, protozoa, and fungi.
3. Interstate movement of microorganisms infectious to livestock/poultry including
bacteria, viruses, protozoa, and fungi, arthropod vectors of livestock/poultry
diseases, and tissues, blood, serum, or cells from known infected
livestock/poultry.

Note: A courtesy letter to the Florida Department of Agriculture and
Consumer Services Division of Animal Industry
http://www.doacs.state.fl.us/ai/contact.shtml is required for possession or use
of any of the State of Florida reportable animal diseases
http://www.doacs.state.fl.us/ai/main/ani diseases main.shtml.


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4. USDA/APHIS Plant Protection & Quarantine Permit
http://www.aphis.usda.gov/ppq/permits/ is needed for import or interstate
movement of plant pests, plant pathogens, biological control agents, bees, plant
pest diagnostic laboratories, soil microbe isolation laboratories, federal noxious
weeds and parasitic plants

5. USDA Biotechnology Regulatory Services Notification or Permit
http://www.aphis.usda.gov/brs/fieldtesting_importation.html for the import,
interstate movement, or field release of genetically engineered plants, arthropods,
and plant-associated microorganisms.

6. Florida Department of Agriculture and Consumer Services Division of Plant
Industry Permit http://www.doacs.state.fl.us/onestop/plt/entnempath.html for the
import into Florida of: arthropods, plant pathogens, nematodes, noxious weeds,
genetically altered (insects, nematodes, plants, plant pests) organisms, biological
control agents.
7. The transfer of Select Infectious Agents and Toxins is also regulated by the
USDA/CDC. Each shipment of listed agents must be registered with the
USDA/CDC through a responsible facility official at both the shipping and
receiving entities. Please contact EH&S Biosafety before sending or
requesting any Select Agents. The current list of Select Agents and Toxins
can be found on in this manual or on our website at
http://www.ehs.ufl.edu/Bio/select.htm.

8. Export of Etiologic Agents of Humans, Animals, Plants and Related Materials
is regulated by the US Department of Commerce, Dept. of State, and Dept. of
the Treasury. A wide variety of etiologic agents of human, plant and animal
diseases, including genetic material, and products which might be used for
culture or production biological agents, will require an export license.
Information may be obtained by calling the Biosafety Office at 352-392-1591.
Export to certain countries is prohibited.


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Biological Safety Cabinets
The following is excerpted from Primary Containment for Biohazards: Selection, Installation and Use of
Biological Safety Cabinets, 1995, Centers for Disease Control & Prevention/National Institutes of Health

This chapter presents information on the
selection, function, and use of biological
safety cabinets (BSCs), which are the
primary means of containment
developed for working safely with
infectious microorganisms. BSCs are
designed to provide personnel,
environment, and product protection
when appropriate practices and
procedures are followed.1

Three kinds of biological safety
cabinets, designated as Class I, II, and -
III, have been developed to meet
varying research and clinical needs.

Biological safety cabinets use high efficiency particulate air (HEPA) filters in their
exhaust and/or supply systems.

Biological Safety Cabinets (BSCs)
The similarities and differences in protection offered by the various classes of biosafety
cabinets are reflected in Table 1.
The Class I BSC
This type of cabinet is not for aseptic or sterile technique. The Class I BSC provides
personnel and environmental protection, but no product protection. It is similar in air
movement to a chemical fume hood, but has a HEPA filter in the exhaust system to
protect the environment.
The Class II BSC
The Class II (Types A and B) biological safety cabinets provide personnel,
environmental, and product protection. Air flow is drawn around the operator into the
front grille of the cabinet, which provides personnel protection. In addition, the
downward laminar flow of HEPA-filtered air provides product protection by minimizing
the chance of cross-contamination along the work surface of the cabinet. Because
cabinet air has passed through the exhaust HEPA filter, it is contaminant-free
(environmental protection), and may be recirculated back into the laboratory (Type A
BSC) or ducted out of the building (Type B BSC).
The Class II, Type A BSC
An unducted Class II Type A BSC is not to be used for work involving volatile or toxic
chemicals. The buildup of chemical vapors in the cabinet (by recirculated air) and in the


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laboratory (from exhaust air) could create health and safety hazards. Generally, BSCs are
not for use with chemicals. Small quantities of chemicals and chemotherapeutic agents
may be used in ducted BSCs.

It is possible to duct the exhaust from a Type A cabinet out of the building. However, it
must be done in a manner that does not alter the balance of the cabinet exhaust system,
thereby disturbing the internal cabinet air flow. The typical method of ducting a Type A
cabinet is to use a "thimble," or canopy unit, which maintains a small opening (usually 1
inch) around the cabinet exhaust filter housing. The volume of the exhaust must be
sufficient to maintain the flow of room air into the space between the thimble unit and the
filter housing (contact manufacturers for any additional specifications). The thimble
must be removable or be designed to allow for operational testing of the cabinet. The
performance of a cabinet with this exhaust configuration is unaffected by fluctuations in
the building exhaust system.

"Hard-ducting" (i.e., direct connection) of Class II Type A cabinets to the building
exhaust system is not recommended. The building exhaust system must be precisely
matched to the airflow from the cabinet in both volume and static pressure. However,
fluctuations in air volume and pressure that are common to all building exhaust systems
make it difficult, if not impossible, to match the airflow requirements of the cabinet.
The Class II, Type B1 BSC
Some biomedical research requires the use of small quantities of certain hazardous
chemicals, such as carcinogens. The powdered form of these carcinogens should be
weighed or manipulated in a chemical fume hood or a static-air glove box. Carcinogens
used in cell culture or microbial systems require both biological and chemical
containment.

Type B1 cabinets must be hard-ducted to their own dedicated exhaust system. As
indicated earlier, blowers on laboratory exhaust systems should be located at the terminal
end of the ductwork. A failure in the building exhaust system may not be apparent to the
user, as the supply blowers in the cabinet will continue to operate. A pressure-
independent monitor should be installed to sound an alarm and shut off the BSC supply
fan, should failure in exhaust airflow occur. Since all cabinet manufacturers do not
supply this feature, it is prudent to install a sensor in the exhaust system as necessary. To
maintain critical operations, laboratories using Type B BSCs should connect the exhaust
blower to the emergency power supply.
The Class II, Type B2 BSC
This BSC is a total-exhaust cabinet; no air is recirculated within it. This cabinet
provides simultaneous primary biological and chemical containment.

Should the building or cabinet exhaust fail, the cabinet will be pressurized, resulting in a
flow of air from the work area back into the laboratory. Cabinets built since the early
1980's usually have an interlock system installed by the manufacturer to prevent the
supply blower from operating whenever the exhaust flow is insufficient. Presence of


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such an interlock system should be verified; systems can be retrofitted if necessary. A
pressure-independent device should monitor exhaust air movement.
The Class II, Type B3 BSC
This BSC is a ducted Type A cabinet. All positive pressure contaminated plenums within
the cabinet are surrounded by a negative air pressure plenum. Thus, leakage in a
contaminated plenum will be into the cabinet and not into the environment.
The Class III BSC
The Class III BSC was designed for work with biosafety level 4 microbiological agents,
and provides maximum protection to the environment and the worker. It is a gas-tight
enclosure with a non-opening view window.

Long, heavy-duty rubber gloves are attached in a gas-tight manner to ports in the cabinet
and allow for manipulation of the materials isolated inside. Although these gloves
restrict movement, they prevent the user's direct contact with the hazardous materials.
The trade-off is clearly on the side of maximizing personal safety. Depending on the
design of the cabinet, the supply HEPA filter provides particulate-free, albeit somewhat
turbulent, airflow within the work environment.

Horizontal Laminar Flow "Clean Bench"
Horizontal laminar flow clean air benches are NOT BSCs. They discharge HEPA-filtered
air across the work surface and toward the user. These devices only provide product
protection. They can be used for certain clean activities, such as the dust-free assembly
of sterile equipment or electronic devices. These benches should never be used when
handling cell culture materials or drug formulations, or when manipulating potentially
infectious materials. The worker can be exposed to materials (including proteinaceous
antigens) being manipulated on the clean bench, which may cause hypersensitivity.
Horizontal clean air benches should never be used as a substitute for a biological safety
cabinet in research, biomedical or veterinary laboratories and/or applications.

Vertical Laminar Flow "Clean Bench"
Vertical laminar flow clean benches also are NOT BSCs. They may be useful, for
example, in hospital pharmacies when a clean area is needed for preparation of
intravenous drugs. While these units generally have a sash, the air is usually discharged
into the room under the sash, resulting in the same potential problems as the horizontal
laminar flow clean benches.

Operations within a Class II BSC

Laboratory Hazards
Many common procedures conducted in BSCs may create splatter or aerosols. Good
microbiological techniques should always be used when working in a biological safety
cabinet. For example, techniques to reduce splatter and aerosol generation will minimize
the potential for exposure to personnel from infectious materials manipulated within the


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cabinet. Class II cabinets are designed so that horizontally nebulized spores will be
captured by the downward flowing cabinet air within fourteen inches of travel. Therefore,
as a general rule of thumb, keeping clean materials at least one foot away from aerosol-
generating activities will minimize the potential for cross-contamination.

The general workflow should be from "clean to contaminated (dirty)." Materials and
supplies should be placed in such a way as to limit the movement of "dirty" items over
"clean" ones.

Several measures can be taken to reduce the chance for cross-contamination when
working in a BSC. Work at least 6" back from the front edge and never cover the front
grill. Opened tubes or bottles should not be held in a vertical position. Investigators
working with Petri dishes and tissue culture plates should hold the lid above the open
sterile surface to minimize direct impact of downward air. Bottle or tube caps should not
be placed on the toweling. Items should be recapped or covered as soon as possible.

Open flames are not permitted in the near microbe-free environment of a biological
safety cabinet. On an open bench, flaming the neck of a culture vessel will create an
upward air current that prevents microorganisms from falling into the tube or flask. An
open flame in a BSC, however, creates turbulence that disrupts the pattern of air supplied
to the work surface. When deemed absolutely necessary, touch-plate microburners
equipped with a pilot light to provide a flame on demand may be used. Internal cabinet
air disturbance and heat buildup will be minimized. The burner must be turned off when
work is completed. Small electric "furnaces" are available for decontaminating
bacteriological loops and needles and are preferable to an open flame inside the BSC.
Disposable sterile loops should be used to eliminate the need for heat or flame.

Aspirator bottles or suction flasks should be connected to an overflow collection flask
containing appropriate disinfectant, and to an in-line HEPA or equivalent filter. This
combination will provide protection to the central building vacuum system or vacuum
pump, as well as to the personnel who service this equipment. Inactivation of aspirated
materials can be accomplished by placing sufficient chemical decontamination solution
into the flask to kill the microorganisms as they are collected. Once inactivation occurs,
liquid materials can be disposed of appropriately as noninfectious waste.

Investigators must determine the appropriate method of decontaminating materials that
will be removed from the BSC at the conclusion of the work. When chemical means are
appropriate, suitable liquid disinfectant should be placed into the discard pan before work
begins. Items should be introduced into the pan with minimum splatter, and allowed
appropriate contact time as per manufacturer's instructions. Alternatively, liquids can be
autoclaved prior to disposal. Contaminated items should be placed into a biohazard bag
or discard tray inside the BSC. Water should be added to the bag or tray prior to
autoclaving.

When a steam autoclave is to be used, contaminated materials should be placed into a
biohazard bag or discard pan containing enough water to ensure steam generation during


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the autoclave cycle. The bag should be taped shut or the discard pan should be covered
in the BSC prior to removal to the autoclave. The bag should be transported and
autoclaved in a leak-proof tray or pan.

Decontamination

Surface Decontamination
All containers and equipment should be surface decontaminated and removed from the
cabinet when work is completed. At the end of the workday, the final surface
decontamination of the cabinet should include a wipe-down of the work surface, the
cabinet's sides and back, and the interior of the glass. If necessary, the cabinet should
also be monitored for radioactivity and decontaminated when necessary. Investigators
should remove their gloves and gowns and wash their hands as the final step in safe
microbiological practices.

Small spills within the BSC can be handled immediately by removing the contaminated
absorbent paper toweling and placing it into the biohazard bag. Any splatter onto items
within the cabinet, as well as the cabinet interior, should be immediately wiped with a
towel dampened with decontaminating solution. Gloves should be changed after the work
surface is decontaminated and before placing clean absorbent toweling in the cabinet.
Hands should be washed whenever gloves are changed or removed.

Spills large enough to result in liquids flowing through the front or rear grilles require
more extensive decontamination. All items within the cabinet should be surface
decontaminated and removed. After ensuring that the drain valve is closed,
decontaminating solution can be poured onto the work surface and through the grille(s)
into the drain pan.

Thirty minutes is generally considered an appropriate contact time for decontamination,
but this varies with the disinfectant and the microbiological agent. Manufacturer's
directions should be followed. The spilled fluid and disinfectant solution on the work
surface should be absorbed with paper towels and discarded into a biohazard bag. The
drain pan should be emptied into a collection vessel containing disinfectant. A flexible
tube should be attached to the drain valve and be of sufficient length to allow the open
end to be submerged in the disinfectant within the collection vessel. This procedure
serves to minimize aerosol generation. The drain pan should be flushed with water and
the drain tube removed.

Should the spilled liquid contain radioactive material, a similar procedure can be
followed. Radiation safety personnel should be contacted for specific instructions (392-
7359).
Gas Decontamination
BSCs that have been used for work involving infectious materials must be
decontaminated before HEPA filters are changed, internal repair work is done or before a
BSC is relocated. The most common decontamination method uses formaldehyde gas,


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although more recently, hydrogen peroxide vapor has been used successfully. This
environmentally benign vapor is useful in decontaminating HEPA filters, isolation
chambers, and centrifuge enclosures. Call the Biosafety Office (392-1591) for the current
BSC vendor who does the gas decontaminations and certifications. All BSCs must be re-
certified following any gas decontamination, maintenance or relocation.

Engineering Requirements

Ultraviolet Lamps
Ultraviolet (UV) lamps are not required in BSCs. If installed, UV lamps must be cleaned
weekly to remove any dust and dirt that may block the germicidal effectiveness of the
ultraviolet light. The lamps should be checked periodically with a meter to ensure that
the appropriate intensity of UV light is being emitted. UV lamps must be turned off
when the room is occupied to protect eyes and skin from UV exposure, which can burn
the cornea and cause skin cancer. Do not depend on UV lamps to disinfect the area.
BSC Placement
The ideal location for the biological safety cabinet is remote from the entry (e.g., the rear
of the laboratory away from traffic), since people walking parallel to the face of a BSC
can disrupt the air curtain. The air curtain created at the front of the cabinet is quite
fragile, amounting to a nominal inward and downward velocity of 1 mph. Open
windows, air supply registers, or laboratory equipment that creates air movement (e.g.,
centrifuges, and vacuum pumps) should not be located near the BSC. Similarly,
chemical fume hoods must not be located close to BSCs.
HEPA Filters
HEPA filters, whether part of a building exhaust system or part of a cabinet, will require
replacement when they become so loaded that sufficient air flow can no longer be
maintained. Filters must be decontaminated before removal.
Certification of BSCs
All BSCs must be certified (according to a National Sanitation Foundation standard)
annually according to UF policy. Please contact the EH&S Biological Safety Office for
the name and phone number of the current contractor performing this service. Prices and
quality vary widely, so only BSO-approved contractors may be used.


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Emergency Procedures/Telephone Numbers

In the event of a major disaster affecting the campus, the UF Homepage http://www.ufl.edu/ is the
official source of UF emergency related information.

Emergency Phone Numbers


Normal
Business
Hours


Evenings & Weekends


Fire/Police/Medical Emergency
Note: callers should be able to accurately state
their location to emergency responders. Write the
location/address of the lab here:




Florida Poison Information Center

Suspected Gas Leak
Needlestick/Sharps Injury
Chemical Spill
http://www.ehs.ufl.edu/RiskMqmt/emerqncy/chemspill.htm
Radiation Spill
http://www.ehs.ufl.edu/Rad/RCGuide/rcq2.htm#21X
Biological Spill
http://www.ehs.ufl.edu/RiskMqmt/emerqncy/biospill.htm
Physical Plant Trouble Desk*
HSC Physical Plant Trouble Desk
IFAS Facilities Operations/Maintenance


911
Note that you
may need to
dial 9 first to
get an outside
line.


911
Note that you may need to dial 9 first to get
an outside line.


1-800-222-
00-222- 1-800-222-1222
1222
392-1121 392-1121
1-866-477-6824
392-8400 or 392111
392-1111
1591
392-7359 or 392111
392-1111
1589

392-1591 392-1111


392-1121
392-4411
392-1984


* 24-hour building/maintenance repair hotline.

Other Contacts
PI or lab Supervisor home number and/or cell numberss:
(fill in)


392-1121
392-4411
392-1121


Karen Gillis, UF BSO, 392-1591


Mark Yanchisin, UF lab safety Coordinator, 392-1591

Phil Collis, Assoc. Director of EH&S, 392-1591

Student Health Care Center 352-392-1161

Alachua County Emergency Management 352-264-6500

Red Cross, Alachua County Headquarters 352-376-4669


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General Information


It is important that persons handling biological agents have the proper training and
experience to work with these materials safely. This includes how to handle an accidental
release, exposure, injury, or evidence of agent theft or loss.
Personnel should know what constitutes a potential exposure or release and
report this to the PI or lab supervisor immediately. The PI or lab supervisor
must then report this to the biosafety office and, as appropriate, to medical care
providers.
All personnel must also be familiar with symptoms of disease or illness
associated with the materials they handle so that a previously undetected
exposure can be discovered and measures taken to prevent further exposures.
Time and situation permitting, contain and secure all biological agents during
any and all emergencies. Agent storage areas shall remain locked.
Know ahead of time the location and operation of emergency equipment such
as the eyewash and safety shower, first aid kit, chemical and biological spill kits,
emergency numbers, fire extinguishers, emergency exits.
PIs or lab supervisors should periodically review this information with lab staff/
students and display this information in the laboratory.
When possible, lab staff, PI/lab supervisor should meet and escort emergency
personnel on site.

Medical Emergency
1) Remain calm.
2) Initiate lifesaving measures, as required.
3) Call for EMERGENCY RESPONSE MEDICAL EMERGENCY 9-1-1.
4) Calling 911 automatically alerts the UF campus Police.
5) Do not remove injured person unless there is a danger of further harm.
6) Keep injured person warm.
7) Notify the PI/ supervisor.

Accidental injection, cuts, skin exposures
1) Remove protective clothing or PPE; place it in biowaste or red bag if contaminated.
2) Wash hands.
3) Wash the effected part. Use soap if available, but avoid strong chemical disinfectants
that can damage skin, e.g. bleach.
4) Apply an appropriate disinfectant from the first aid kit (e.g. antibiotic ointment).
5) Notify the PI or lab supervisor and inform them of the circumstances of the injury,
including what was being handled at the time.
6) If medical treatment is needed:
a) During working hours go to the UF Student Health Center (SHCC):


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i) Main Campus (Infirmary) location:
Fletcher Drive next to the Florida Gym and the Racquet Club, 392-1161
ii) UF Health Science Center Satellite location:
Second floor of the Dental Towers of the J. Hillis Miller Health Science
Center. Take a Dental Tower elevator to the second floor, turn left, and
straight ahead is SHCC@Shands in Room D2-49, phone 392-0627

b) After hours or on weekends, seek treatment at the nearest emergency room.
c) The PI or supervisor must notify the Biosafety office of the incident.

7) If exposed to a Blood Borne Pathogen:
a) Wash the area thoroughly with soap & water
b) In Gainesville, call 1-866-477-6824, the Needle Stick Hotline.
c) In Jacksonville, 7:00 4:00, Go to Employee Health Suite 505 in Tower 1. Other
hours, go to the ER
d) Other areas: Go to nearest medical facility
e) Get immediate medical attention (1-2 hr max)
f) Notify supervisor
g) Allow Medical to follow up with the appropriate testing & the required written
opinion.

Splashes to face and eyes
1) Go to the eyewash station and activate it
2) Rinse face/mouth/nose/eyes
3) Eyes should be flushed for at least 15 minutes.
4) Forcibly hold eye open to ensure effective rinsing behind eyelids.
5) Have injured worker move eye side-to-side and up-down during rinsing.
6) Remove contact lenses.
7) Always obtain medical attention for a hazardous material splash to the eye.
8) Place contaminated clothing in a red biohazard bag for decontamination.
9) Watch for symptoms of exposure or delayed onset effects.
10) Report incident to PI/supervisor and Biological Safety Office (392-1591).

Accidental ingestion
1) Seek medical treatment:
a) During working hours go to the UF Student Health Center (SHCC):
i) Main Campus (Infirmary) location: Fletcher Drive next to the Florida Gym
and the Racquet Club, 392-1161
ii) UF Health Science Center Satellite location: Second floor of the Dental
Towers of the J. Hillis Miller Health Science Center. Take a Dental Tower
elevator to the second floor, turn left, and straight ahead is SHCC@Shands in
Room D2-49, phone 392-0627


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b) After hours or on weekends, seek treatment at the nearest emergency room.

2) The PI or supervisor must notify the Biosafety office of the incident.


Animal bites and scratches
1) For small wounds allow to bleed freely. If necessary, control bleeding by applying
direct pressure with a sterile gauze or bandage.
2) Immediately wash with copious quantities of soap and water. Wash for at least 15
minutes. A chlorhexidine soap such as Nolvason is recommended. Povidone iodine
or Betadine surgical soap may be used but is more likely to cause skin irritation and
cellular damage. If eyes or mucous membranes are exposed, irrigate the area for at
least 15 minutes with water.
3) Seek medical treatment:
a) During working hours go to the UF Student Health Center (SHCC):
i) Main Campus (Infirmary) location: Fletcher Drive next to the Florida Gym
and the Racquet Club, 392-1161
ii) UF Health Science Center Satellite location: Second floor of the Dental
Towers of the J. Hillis Miller Health Science Center. Take a Dental Tower
elevator to the second floor, turn left, and straight ahead is SHCC@Shands in
Room D2-49, phone 392-0627
b) After hours or on weekends, seek treatment at the nearest emergency room.
4) The PI or supervisor must notify the Biosafety office of the incident.
5) If the bite or scratch is from a non-human primate, contact the following
physician/specialists regarding Monkey B virus (Herpesvirus Simiae, CHV-1)
exposure:
a) Dr. Reuben Ramphal, Work phone: 352 392-4058, Pager: 352-413-7837
b) Dr. Kenneth Rand, Work phone: 352 392-5621, Pager: 888-553-2503
c) The physicians will evaluate the injury and may decide to culture the wound for
B-virus (Herpesvirus simiae) or collect blood for a baseline titer against B-virus,
or use prescription drugs for preventative therapy. The physician directing the
care of the patient will contact the Director of Animal Care Services for
instructions regarding the need for cultures or serology from the monkey
inflicting the injury upon the patient.
d) Symptoms suggestive of B virus infection should be reported immediately to the
medical consultant. When the possibility of B virus illness is seriously
entertained, appropriate diagnostic studies should be performed and specific
antiviral therapy should be instituted. The physician may wish to consult the Viral
Exanthems and Herpesvirus Branch, Division of Viral and Rickettsial Diseases,
CDC (Dr. Scott Schmitt, (404) 639-0066 or cell 404-725-5652 or Terri Hyde
(404) 639-2696, for laboratory assistance, the National B Virus Resource Center
at GSU, (Dr. Julia Hilliard, (404) 651-0808).
e) The above procedures also apply to the employees of private contractors in non-
human primate facilities.


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Break in/Security Breach
1. Call 352-392-1111 for University police department.
2. Notify PI/lab supervisor and the Biosafety Office of the break in.
3. Escort University Police personnel at the scene.
4. The PI and Biosafety Office will conduct an inventory check and report results to
appropriate authorities as needed.

Handling Biological Spills
Advance preparation for management of a spill is essential. A "bio spill kit" should be
available and contain the following:
* Forceps/scoop for broken glass/sharps
* Paper towels or absorbent material
* Disinfectant
* Respirators, if necessary
* Latex or nitrile gloves and safety glasses
* Red bag

Spill in the biosafety cabinet
1) Leave the cabinet on/running to prevent escape of contaminants from the cabinet.
2) Cover the area with paper towels or other absorbent material.
3) Pour appropriate disinfectant (e.g. a fresh 1:10 dilution of household bleach, 0.5%
sodium hypochlorite) over the spill as a disinfectant solution to inactivate
biohazardous material. If necessary, sufficient disinfectant solution shall be used to
ensure that the drain pans and catch basins below the work surface contain
disinfectant. Disinfect under the front exhaust grill if needed. Walls and equipment
in the biological safety cabinet that may have been splashed shall be wiped with
disinfectant.
4) Let disinfectant solution sit for 30 minutes.
5) Pick up absorbent materials and wipe up excess disinfectant solution.
6) Place material in biohazard bag.
7) Use tweezers, forceps, or tongs to pick up broken sharps and place in a sharps
container.
8) Rinse all disinfected areas with water and allow to dry.
9) Follow with 70% ethanol.

Spill in the centrifuge

1) Allow aerosols to settle for 30 minutes before attempting to clean up the spill. Keep
the centrifuge closed during this time. Post a sign on the centrifuge so others don't
try to open it.
2) Gently open the centrifuge to prevent re-aerosolization.
3) Place absorbent materials in the chamber and pour a fresh 1:10 dilution of household
bleach (0.5% sodium hypochlorite) over them. Let sit 30 minutes.


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4) Carefully remove carriers to a tub containing a fresh 1:10 dilution of household
bleach (0.5% sodium hypochlorite). Soak 30 minutes.
5) Wipe the interior and lid of the centrifuge with a fresh 1:10 dilution of household
bleach (0.5% sodium hypochlorite).
6) Use tweezers, forceps, or tongs to pick up broken sharps place in a sharps
container.
7) Wipe all areas with plenty of water to prevent corrosion. Dry and follow with 70%
ethanol.


Spill inside the laboratory


1) Notify room occupants of the spill.
2) All persons should leave the laboratory immediately.
3) Close the door.
4) If clothing is known (or suspected) to be contaminated, remove the clothing with
care, folding the contaminated area inward. Place the clothing into a red bag for
autoclaving.
5) Wash all potentially contaminated body areas as well as the arms, face and hands.
Shower if necessary.
6) Any exposed persons should seek medical advice or treatment:
a) During working hours go to the UF Student Health Center (SHCC):
i) Main Campus (Infirmary) location: Fletcher Drive next to the Florida Gym
and the Racquet Club, 392-1161
ii) UF Health Science Center Satellite location: Second floor of the Dental
Towers of the J. Hillis Miller Health Science Center. Take a Dental Tower
elevator to the second floor, turn left, and straight ahead is SHCC@Shands in
Room D2-49, phone 392-0627
b) After hours or on weekends, seek treatment at the nearest emergency room.
7) Post a sign on the door to keep people out.
8) No one should enter for approximately 30 minutes so that aerosols can be cleared by
the ventilation system or allowed to settle.
9) During this time, notify the PI or lab supervisor. Notify the biosafety office for large
spills, those you are uncomfortable handling, those involving BSL-3 agents, or those
involving select agents.
10) Protective clothing should be worn when entering the laboratory to clean the spill
area. Latex or nitrile gloves, autoclavable, or disposable footwear, safety glasses, and
an outer garment. If you have been issued an N95 respirator to work with this agent,
put that on.
11) Take the "bio spill kit" into the laboratory room, and place paper towels, spill
pillows, or other absorbent materials around and on the spill. If the spill was on the
floor, do not use a surgical gown that may trail on the floor when bending down.
12) Carefully pour a fresh 1:10 dilution of household bleach (0.5% sodium
hypochlorite) over the spill as a disinfectant solution; avoid splashing and work
from the outside towards the center.
13) Let disinfectant solution sit for 30 minutes.


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14) Use paper or cloth towels to wipe up the disinfectant and spill, working toward the
center of the spill. Discard all towels and other clean up materials into a discard
container/red bag as they are used.
15) Wipe the outside of the discard containers, especially the bottom, with a towel
soaked in a disinfectant.
16) Place the discard container and other materials in an autoclave and sterilize.
17) Remove shoes or shoe covers, outer clothing, respirator, and gloves and sterilize by
autoclaving.
18) Wash hands, arms and face; shower if necessary.
19) If gaseous decontamination of the room is required, contact the Biosafety Office.

Spill outside the laboratory


Safe transport of biohazardous material outside the laboratory is essential. Materials
should be packaged securely (triple contained in unbreakable containers) to avoid such
spills. In addition, the person transporting the material should be knowledgeable about
the hazards of the material and how to respond to a spill. In the event of a spill outside
the lab:

1) Clear area of all personnel and keep them out of the spill
2) Have someone call the University of Florida Police Dept., 392-1111 and the
Biosafety Office 392-1591, for help.
3) If clothing is known (or suspected) to be contaminated, remove the clothing with
care, folding the contaminated area inward. Place the clothing into a bag for
autoclaving.
4) Wash all potentially contaminated body areas as well as the arms, face and hands.
Shower if necessary.
5) Any exposed persons should seek medical advice or treatment:
a) During working hours go to the UF Student Health Center (SHCC):
i) Main Campus (Infirmary) location: Fletcher Drive next to the Florida Gym
and the Racquet Club, 392-1161
ii) UF Health Science Center Satellite location: Second floor of the Dental
Towers of the J. Hillis Miller Health Science Center. Take a Dental Tower
elevator to the second floor, turn left, and straight ahead is SHCC@Shands in
Room D2-49, phone 392-0627
b) After hours or on weekends, seek treatment at the nearest emergency room.
6) Spill will be cleaned up by lab staff (if a spill kit is available) or by EH&S staff.
7) Protective clothing should be worn to clean the spill area. Latex or nitrile gloves,
autoclavable, or disposable footwear, safety glasses, and an outer garment. If you
have been issued an N95 respirator to work with this agent, put that on.
8) Take the "bio spill kit" to the area, and place paper towels, spill pillows, or other
absorbent materials around and on the spill. If the spill was on the floor, do not use
a surgical gown that may trail on the floor when bending down.


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9) Carefully pour a fresh 1:10 dilution of household bleach (0.5% sodium
hypochlorite) over the spill as a disinfectant solution; avoid splashing and work
from the outside towards the center.
10) Let disinfectant solution sit for 30 minutes.
11) Use paper or cloth towels to wipe up the disinfectant and spill, working toward the
center of the spill. Discard all towels and other clean up materials into a discard
container/red bag as they are used.
12) Wipe the outside of the discard containers, especially the bottom, with a towel
soaked in a disinfectant.
13) Place the discard container and other materials in an autoclave and sterilize.
14) Remove shoes or shoe covers, outer clothing, respirator, and gloves and sterilize by
autoclaving.
15) Wash hands, arms and face; shower if necessary.

Fire Safety


Predetermine two means of egress from your normal workplace.
Learn the location of the nearest fire alarm pull station and portable fire extinguisher.

If you discover a fire in a University of Florida building, do the following:
1) Pull the fire alarm and call 911.
2) Fire alarm pull stations are normally located near each exit. If the building is not
equipped with a fire alarm system, notify other occupants as you exit the building.
3) Do not attempt to fight the fire with portable fire extinguishers or fire hoses unless
the fire is small and you have been trained in their proper use. DO NOT PUT YOUR
LIFE IN DANGER WHILE ATTEMPTING TO CONTROL A FIRE. When in
doubt, evacuate.
4) Remain calm while talking to the operator. Be prepared to answer several questions
as to location, size of fire, your name, number of persons in building (if known) and
any injuries. Remain on the line until the operator is finished.
5) Meet fire or police personnel when they arrive at the building. Stand by to answer any
questions they may have concerning the fire.
6) Once out of the building DO NOT RE-ENTER THE BUILDING FOR ANY
REASON, unless emergency personnel have given the "ALL CLEAR" signal.

If the fire is INSIDE your room:
Leave your room and close the door.
Pull the fire alarm and call 911.
Or, if the fire is small and you have been trained to use a fire extinguisher, attempt to put
it out. (Again, DO NOT PUT YOUR LIFE IN DANGER WHILE ATTEMPTING TO
CONTROL A FIRE. When in doubt, evacuate.)
Remember the acronym "PASS"
PASS =
P Pull the pin.
A Aim at the base of the fire.


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S Squeeze the trigger.
S Sweep the nozzle from side to side.


If the fire is NOT in your room, but in a room you must pass through to get out or
exit:
1) With your hands, test the door for heat before opening.
2) If the door is HOT:
a) Stay in your room or lab.
b) Phone for help.
c) Stay calm.
d) Seal cracks with wet towels.
e) Wait for help.
3) If the door is COOL:
a) Take your room key.
b) Open the door slowly.
c) WALK to the nearest exit and leave the building.
d) If the exit is unsafe, return to the room and remain there.
e) If the hall is smoky, stay low or crawl out on your hands and knees.

DO NOT USE THE ELEVATOR !!!


Workplace Violence:

All threats and other inappropriate behavior that create an immediate concern for safety
should be reported immediately to the University Police Department (UPD) at (352) 392-
111 lor local law enforcement if off campus. You may also dial 911, but remember, you
must first dial 9 to get an outside line.

Examples of behavior requiring a call to authorities include:
Direct or veiled threats
Written sexual or violent notes intimidation verbally or physically
Carries a weapon (Florida Statutes and University Policy prohibit firearms and certain
other articles that could be weapons on state property)
Makes suicidal comments or threats
Involved in fights or assaults
Stalks co-workers or their family
See the UF Workplace violence policy at
http://www.hr.ufl.edu/policies/workviolencepolicy.pdf



Hurricane:


1) All biological agents shall be secured and contained in preparation for a hurricane.


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2) See the checklist to prepare lab areas for an impending hurricane:
http://www.ehs.ufl.edu/disasterplan/LabPrep.rtf
3) The UF homepage is the official source of UF Information. Official emergency
information for Alachua County is broadcast on
i) Radio: WUFT-FM / WJUF-FM / WLUF-LP / WRUF / AM850 / Rock 104
ii) Television: WUFT-TV

iii) Additional information regarding safety and preparedness may be
found at: www.ufl.edu/emerg.html



Tornadoes and other natural disasters:


Tornado Watch conditions are favorable for the formation of tornados.
Tornado Warning indicates that a tornado has been sighted or is indicated on radar.
There is generally little or no warning given at the approach of a tornado. In the event of
a tornado:
1) IMMEDIATELY seek shelter, preferably in the hallway of a main office building, an
interior bathroom or an interior closet.
2) AVOID windows; flying debris can kill. Protect yourself by getting under a heavy
desk or table. Also, remember to protect your head.
3) If time permits, open at least two windows to provide complete ventilation for each
building.


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3 -- Programs


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Animal Contact Medical Monitoring Program
The University of Florida Animal Contact Medical Monitoring Program
grew out of the recommendations of the American Association for the
Accreditation of Laboratory Animal Care (AAALAC) reviewers.
According to the US Public Health Service, an occupational health
program is required for institutions that employ personnel who have animal
contact.

The UF program is designed to protect our employees, students, and volunteers having
animal contact from occupational exposure to conditions that may result in animal related
illnesses. The requirements of the program are based upon those outlined in the Public
Health Services document, Guidefor the Care and Use of Laboratory Animals, published
by the National Research Council. The current UF program requirements were revised
and approved, and put into place May 2006.

Individuals who will be working with animals or who will be working in proximity to
animals are required to participate in the medical monitoring program. They are provided
with animal contact medical monitoring information and immunizations relating to their
animal contact as part of their pre-placement health assessment.

In addition, the Institutional Animal Care and Use Committee (IACUC) verifies that all
personnel listed on new and continuing projects are registered with the UF Animal
Contact Medical Monitoring Program. The IACUC committee notifies the PI of any
personnel who are not thus cleared for animal contact. Principal Investigators are
responsible for ensuring that all personnel (including employees, students, colleagues,
collaborators, and volunteers) involved with their IACUC-approved project are given
program information. Investigators who do not respond to requests for registration may
have their approval rescinded by the IACUC.

Individuals that have animal contact must participate in a risk assessment that includes
contact information, health questionnaire and health assessment (physical examination,
medical history, blood serum banking) based upon the type of animal contact, and
immunizations as needed. This risk assessment is provided at no cost to the employee.

Short-term visitors to UF from other institutions must provide documentation of
participation in their home institution's animal contact program or they must register in
the UF Animal Contact Program. Individuals involved in isolated one-time, non-recurrent
exposures shall be informed of potential dangers and medical precautions, but are not
required to participate in the program. The primary responsible party (principal
investigator, research director, student research coordinator, etc.) shall be responsible for
assuring compliance with the notification requirements for these individuals.


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Elements of the Program


1. Risk Assessment

A Risk Assessment form is required to be completed by everyone working with
animals at UF or entering UF animal facilities. This form includes contact
information and a health questionnaire that will be evaluated by UF's
Occupational Medicine Physician or Licensed Health Care Professionals to assess
the risk of exposure and determine whether additional information and/or
interaction is necessary.

A Renewal Risk Assessment form is required every three years or when any new
species is contacted. This update will allow UF's Occupational Medicine
Physician or Licensed Health Care Professionals to evaluate and, if necessary,
address potential health risks to you resulting from a change to your health status
or changes to the type of animal exposure.

2. Specific requirements

Tetanus Immunization within 10 years All participants

Rabies Immunization Series/Booster or Positive Titer within 2 years All
individuals handling unvaccinated carnivores or their tissue

Respirator Clearance and Fit Test All individuals required by the Q-Fever Policy
or as medically necessary to prevent allergic reactions

Serum Banking All individuals who work with non-human-primates, all who
handle blood from alligators or birds housed outdoors, all who are required by the
Q-Fever Policy and all pre-menopausal females who handle cats or cat waste.

TB Screening within 12 months All individuals who enter any room with non-
human primates.

Medical consultation As determined by the Occupational Medicine Physician.
Examples are individuals with chronic disease, work-related injuries or illness,
environmental or animal allergies.

3. Exemptions from the program

Individuals working on projects that involve observation of birds or other animals
outdoors/in their natural habitat are exempt from the program.

See http://www.ehs.ufl.edu/Bio/Animal/acweb.htm for more information.


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UF Bloodborne Pathogen Program
In December 1991, OSHA published the final rule covering
occupational exposure to bloodbome pathogens. This was
adopted by the state of Florida and written into the Florida
Administrative Code in January 1993. UF instituted its program
in the spring of 1993.

The rule requires that those who handle human blood or other potentially infectious
(human) materials as part of their job duties participate in an employer-generated
program. This program shall include development and annual review of a site specific
Exposure Control Plan, annual training regarding exposures, offer of hepatitis B
vaccinations free-of-charge, and post-exposure health care services.

Environmental Health and Safety (EH&S) manages this program through the Biological
Safety Office. Each January, EH&S sends program materials and instructions to
department chairs and directors of groups identified as having employees with
occupational exposure. These materials provide instructions and information necessary to
achieve annual training compliance. The BBP program materials and information is
available on our website at http://www.ehs.ufl.edu/bio/BBP/default.htm.

In addition to the program materials, EH&S provides "train-the-trainer" sessions annually.

Hepatitis B vaccinations are given by the Student Health Care Center (SHCC) or, in some
cases, Shands Occupational Health Services, or other providers. Official program medical
records are kept by SHCC. There is a requirement that Shands or other providers send
vaccination and post-exposure records to SHCC for record-keeping purposes. Declination
statements from those that decline the vaccination series are kept by EH&S.

EH&S monitors for UF compliance by requiring training and vaccination documentation
and by confirming BBP participation during the annual laboratory safety survey which is
conducted in each laboratory each year.

For further information, please call the Biological Safety Office at 392-1591.

BBP Exposure Information
In Gainesville, call the Needlestick Hotline 24 hours a
U F Employees and Students day following any exposure.


1-866-477-6824
0 UC H


In Jacksonville, go to the Employee Health Office in
Suite 505, Tower 1, 5th and Jefferson from 7 4:30.
Go to the ER after hours. Phone: 904 244-9576

For exposure incidents more than 1 hour away from
Gainesville, go to the nearest medical facility.


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4 -- Medical Surveillance










HIV Research Laboratory

Occupational Medicine Policy


This policy is designed to protect employees who conduct research
with HIV. Researchers who handle, manipulate, or assay live HIV
cultures are covered under this policy.

Pre-employment
Prior to beginning work in an HIV research lab, an employee shall be given, at no cost to
the employee, the following:

1. A baseline serum sample shall be stored with the UF Student Health Care Center.
This is a requirement to work in the HIV lab.
2. A confidential HIV test shall be offered to the employee. The HIV test is
available at the UF Student Health Care Center. The test must be offered by the
employer, but may be declined by the employee. This test shall include HIV
counseling. Results of the test shall be given to the employee in person in a face-
to-face meeting with a health care professional. No exceptions.
All results of HIV testing shall remain completely confidential and shall be stored
in a separate section in the employee's medical record. At no time, will the
employer or any administrator or official of UF have access to the employee's
confidential record concerning HIV testing.
3. If the HIV researcher also works with human blood or other potentially infected
material, all aspects of the UF Bloodborne Pathogen Program under the OSHA
Bloodborne Pathogen standard shall be implemented for that employee. The
employer shall offer free HBV immunization, annual training, and have a written
exposure control plan in the workplace.

Continuing employment
HIV researchers shall be given the following at no cost to the employee:
1. Annual serum banking shall be offered to the employee, but may be declined.
2. Annual HIV tests shall be offered to the employee, but may be declined.
3. Annual training regarding HIV and post-exposure prophylaxis, including the risks
of chemoprophylaxis, shall be given to the employee. This is a required, not
optional, component.

Post-exposure prophylaxis
An exposure to an HIV culture in a research laboratory is considered a "highest risk
exposure" according to Public Health Service CDC guidelines. The following shall be
implemented:


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