University of Rochester Institutional Biosafety Committee
Checklist for Biosafety Level 3
Information about biosafety assessment and the Institutional Biosafety Committee (IBC) can be found on the
IBC web site http://www.safety.rochester.edu/ibc/
P.I. _________________________________ Dept. ____________________ Phone____________________
Office Location ________________________e-mail______________________________________________
Project or Grant Title_______________________________________________________________________
Award Number ___________________________________________________________________________
UCAR Number____________________________________________________________________________
Technician _____________________________________ Phone ___________________________________
Recombinant DNA and non-mammalian virus vectors (rDNA plasmids, bacteriophages, baculovirus, insect
cells, yeast, lab strains of E. coli)
Host(s) (whole animals or microbes) _____________________________________________________
Non-human Cell Line(s) and Their Origin _________________________________________________
Vector(s)___________________________________________________________________________
Gene(s) ____________________________________________________________________________
Source of DNA (organism) ____________________________________________________________
Room(s) Number(s) where constructs are used______________________________________________
Laboratory Biosafety Level_________________
Animal Biosafety Level _____________________
Mammalian cells, cell lines and unfixed tissues (this section should include information on tissues or cells
which you intend to harvest from vertebrate animals, in the course of your experiments)
Name of Cell(s), Cell Line(s) and unfixed tissues, and their species of origin______________________

Source of cell line (Vendor; Vendor part number)_____________________________________
_____________________________________________________________________________
(see http://www.atcc.org; note that ATCC recommends specific containment levels for all cells)

Is the cell line known to harbor any infectious agent? If so, please identify the agent.
____________________________________________________________________________

Are you using primary human cells, unfixed human tissues or human body fluid?____________
(If the answer is yes, your work must proceed at BSL2 or higher)
Room(s) Number(s) where cells are used______________________________________________
Laboratory Biosafety Level_________________
Animal Biosafety Level _____________________
Biohazards (infectious agents, mammalian virus vectors; for investigators who intend to use mammalian virus
vectors, the necessary IBC registration form should also be completed; see:
http://www.safety.rochester.edu/ibc/ibcvirus.htm.
Name of Agent(s) ____________________________________________________________________
Room(s) Number(s) where biohazards are used______________________________________________
Laboratory Biosafety Level_________________
Animal Biosafety Level _____________________
A summary or abstract must accompany this form for IBC approval.
I understand that the attached forms are subject to approval by the Institutional Biosafety Committee. A site visit
is part of the approval process for biosafety level 3 containment.
If there are any changes to the protocol, I understand that it is my responsibility to notify the IBC Executive
Secretary using the IBC Biosafety Update Form http://www.safety.rochester.edu/ibc/ibcforms.htm.
_________________________________________ ________________________________
Principal Investigator
Date
IBC SIGNOFF
Facilities and procedures have been reviewed and are adequate and consistent with the appropriate guidelines.
_________________________________________
Biosafety Officer
________________________________
Date
_________________________________________
IBC Chair (Required for BSL-3)
________________________________
Date
_________________________________________
IBC UCAR Coordinator (Required for ABSL-2 and 3)
________________________________
Date
_________________________________________
BSL-3 Director (Required for BSL-3)
________________________________
Date
_________________________________________
Full IBC Committee Approval
________________________________
Date
General Questions for Experiments using Recombinant DNA
Circle “Yes” or “No” for the following questions:
Experiments that require RAC review; NIH and IBC approval.
(Yes/No)
1. Deliberate formation of rDNAs containing genes for biosynthesis of toxic molecules.
(Yes/No)
2. Deliberate release into the environment of any organism containing rDNA.
(Yes/No)
3. Deliberate transfer of drug resistance trait to microorganisms such that drug control might
be compromised.
(Yes/No)
4. Deliberate transfer of rDNA into human subjects.
Experiments that require IBC approval before initiation. Circle "Yes" or "No":
(Yes/No)
1. Use of other than a Risk Group 1 agent as host-vector system (see Appendix B of NIH
Guidelines).
(Yes/No)
2. Will you use a Risk Group 2, 3, or 4 viral vector? If so, will:
(Yes/No) A. greater than 2/3 of genome be used?
(Yes/No) B. helper virus or packaging cells be used?
(Yes/No) C. your experiment enhance pathogenicity (e.g., insertion of oncogene, extend
host range)?
(Yes/No)
3. Will whole animals or plants be used as hosts?
(Yes/No)
4. Will experiments involve more than 10 liters of culture?
(Yes/No)
5. Will a deliberate attempt be made to obtain expression of a foreign gene?
If so, what protein/RNA will be produced?
If you require more information, please read the NIH Guidelines. These are available online at:
http://www4.od.nih.gov/oba/rac/guidelines/guidelines.html
Specific Regulations for Use of Recombinant Virus Vector Systems
If you are intending to use recombinant virus vector systems (in vitro or in vivo), the vectors must be registered
with the IBC using our online virus vector registration form.
This form is completed online at: http://www.safety.rochester.edu/ibc/ibcvirus.htm .
Checklist Instructions
To fill out the Checklist for Biosafety Level 3, start with question 1. Please read carefully and check yes or no.
Feel free to add notes of explanation in the margins.
 IBC Approval must be completed PRIOR to initiation of biosafety level 3 experiments.
 All individuals working at BSL-3 containment must be BSL-3 Authorized Users as per the BSL-3
Program. Contact the Biosafety Officer (x5-3014 or jives@safety.rochester.edu) for more information.
 Compliance with the BSL-3 Program and medical surveillance program is required.
 Personal protective equipment (PPE) must be provided to employees, and must be suitable for the
task(s) performed. PPE includes lab coats, gloves, face protection (if appropriate/necessary) and other
protective measures (if appropriate/necessary).
 A plan for containment of sharps should be in place (e.g., use of sharps containers; no re-capping of
needles, etc).
 Changes in the proposed of recombinant DNA materials or biohazards must be documented:
Use in Whole Animals (includes lab work on animal specimens):
Submit UCAR “Sections A and C” to IBC Executive Secretary.
Use in the Laboratory ONLY:
Submit “IBC Biosafety Update Form” to IBC Executive Secretary
OR
Submit a revised Checklist
 Return completed checklist with an ABSTRACT or summary of the research to Janet Ives,
Biosafety Officer, Executive Secretary at Environmental Health & Safety, 300 East River Road,
Room 23.
 If you need help completing this form, contact Janet Ives, Biosafety Officer, Executive Secretary,
jives@safety.rochester.edu, Telephone: 275-3014.
 Biosafety Level 3 Checklist
YES
1. The Principal (Co) Investigator is familiar with the most current “NIH Guidelines
for Research Involving Recombinant DNA Molecules” (NIH Guidelines), the
CDC/NIH “Biosafety in Microbiological and Biomedical Laboratories”, and with the
UR
Biosafety
Level
3
Program.
(Available
on-line
at
http://www.safety.rochester.edu/ibc/ibclinks.htm)
2. The PI reports within 30 days to the IBC and NIH (ORDA) all significant
problems with and violations of the NIH Guidelines and all significant research
related illnesses.
3. The PI immediately reports all laboratory related incidents resulting in personnel
injury or exposure to the Biosafety Officer.
4. The PI controls access to the laboratory and restricts access to persons whose
presence is required for programs or support purposes. The PI is responsible for
assessing each circumstance and determining who may enter or work in the
laboratory. All individuals working at BSL-3 must comply with the BSL-3 Program.
4. The PI has established policies and procedures whereby only persons who have
been advised of the potential biohazard and meet specific entry requirements may
enter the laboratory or animal rooms.
5. A plan for emergencies has been developed and is available for laboratory
employees. Emergencies to be planned for include fire, hazardous material spills
(chemical, biological, radiological), injuries, and exposures. Spill plans must
specifically address the hazards in the laboratory.
6. Lab specific biosafety information is available, and personnel are required to be
familiar with it. Information should be included: general information regarding
biohazardous agent, routes of disease transmission, recommended vaccinations, signs
and symptoms of disease, required personal protective equipment, waste handling
protocol, spill clean up procedures for inside and outside containment equipment
including centrifuges, exposure follow-up procedure, aerosol control procedure, and
general biosafety information (NIH Guidelines and/or CDC/NIH “Biosafety in
Microbiological and Biomedical Laboratories”).
7. The PI ensure proper instruction of laboratory staff in microbiological techniques,
approved protocols, and emergency procedures, and that these instructions are
followed. Documentation of training is kept by the PI (contents, trainer, attendees,
and date).
8. The PI ensured that before working at biosafety level 3 containment personnel
demonstrate proficiency in standard microbiological practices and techniques, and in
the practices and operations specific to the laboratory facility. Documentation of this
training is kept by the PI (contents, trainer, attendees, and date).
9. All persons working with human blood, body fluids, or tissues receive the OSHA
Bloodborne Pathogens Training annually and have been offered the hepatitis B
vaccination series.
10. Laboratory personnel receive appropriate immunizations or tests for the agents
handled or potentially present in the lab.
11. Persons under 16 years of age do not enter the laboratory.
NO
YES
12. BL-3 laboratory doors are closed while experiments are in progress.
13. The universal biohazard sign is posted on all laboratory access doors and is
posted on all units used to store biohazardous materials. The sign includes agents(s),
name and telephone numbers of responsible individuals, and specific entry
requirements.
14. The BL-3 laboratory is separate from areas that are open to unrestricted traffic
flow within the building. Passage through two sets of doors is the basic requirement
for entry into the laboratory from access corridors or other laboratories. Physical
separation may be achieved by a double-door clothes change room (showers may be
included), airlock, or other access facility which requires passage through two sets of
doors before entering the BL-3 laboratory.
15. Access doors to the BL-3 laboratory are self-closing.
16. The interior surfaces of walls, floors, and ceiling are water resistant and smooth
so that they can be easily cleaned. Penetrations in these surfaces are sealed or capable
of be sealed to facilitate decontaminating the area.
17. Bench tops are impervious to water and resistant to acids, alkalis, organic
solvents, and moderate heat.
18. Laboratory furniture is sturdy and spaces between benches, cabinets, and
equipment are accessible for cleaning.
19. The BL-3 laboratory contains a sink for hand washing. The sink is foot, elbow, or
automatically operated and is located near the laboratory exit.
20. The BL-3 laboratory contains a readily available plumbed eyewash station. The
eyewash is activated weekly.
21. Windows in the BL-3 laboratory are closed and sealed.
22. Vacuum lines are protected with HEPA filters and liquid disinfectant traps, or the
equivalent, which are routinely maintained and replaced as needed.
23. A ducted exhaust air ventilation system is provided. This system creates
directional airflow that draws air into the laboratory through the entry area. The
exhaust air is not recirculated to any other areas of the building, is discharged to the
outside, and is dispersed away form occupied areas and air intakes. All exhaust air
from the laboratory is HEPA filtered.
24. Personnel verify that the direction of the airflow (into the laboratory) is proper
and records are kept.
25. If the HEPA filtered exhaust air from a Class II biosafety cabinet is 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.
NO
YES
NO
26. Biosafety cabinets are decontaminated and certified annually.
Certifier: __________________________________________________________________________
Manufacturer: ______________________________ Model: _________________________________
Class II Type: ______________________________ Serial No.: _______________________________
Certifier: __________________________________________________________________________
Manufacturer: ______________________________ Model: _________________________________
Class II Type: ______________________________ Serial No.: _______________________________
27. Continuous flow centrifuges or other equipment that may produce aerosols are
contained in devices that exhaust air through HEPA filters before discharge into
laboratory.
28. All wastes from BL-3 laboratories and animal rooms are appropriately
decontaminated before disposal.
Procedure: ___________________________________________________________
30. An autoclave for decontaminating laboratory wastes is available preferably
within the laboratory.
Autoclave location:___________________________________________________
30. Materials to be decontaminated away from the laboratory are packaged in durable
leak-proof containers, which are closed before removal from the laboratory.
31. Pipetting by mouth is prohibited. Mechanical pipetting devices are used.
32. Eating, drinking, smoking, handling contact lenses, and applying cosmetics are
not permitted in the lab. Persons wearing contacts also wear goggles or a face shield.
Food storage is prohibited in the BL-3 laboratory.
33. Hand washing facilities are available, and persons wash their hands after
handling recombinant DNA or other biohazardous materials and before leaving the
BL-3 laboratory.
34. Other experiments conducted concurrently are performed following BL-3
precautions even if such experiments are approved for a lower biosafety level.
35. Care is taken to minimize the creation of aerosols.
36. Appropriate combinations of personal protective equipment and containment
equipment are used for all activities with organisms containing recombinant DNA or
infectious materials. No work in open vessels is conducted on the bench.
37. Laboratory equipment and work surfaces are decontaminated daily and
immediately following spills of viable material.
Decontaminant :________________ _____________________________________
38. Spills of infectious materials are contained, decontaminated, and clean up by
appropriate professional staff, or other properly trained and equipped to work with
concentrated infectious materials.
YES
39. Equipment is decontaminated before it is sent for repair or maintenance and
packaged for transport in accordance with applicable local, state, or federal
regulations, before removal from facility.
40. Needles and syringes are used only for parenteral injection and fluid aspiration
from animals and diaphragm bottles. Only locking or integral-type syringes are used.
41. Extreme caution is used when handling needles and syringes to avoid
autoinculation and generation of aerosols during use and disposal.
42. Contaminated needles are not sheared, bent, or recapped.
43. Sharps including needles, razors, scalpels, contaminated broken glass and pasteur
pipettes are disposed of in a sharps shelter.
44. Wrap around or closed front laboratory clothing, shoe covers, head cover, and
gloves are worn at all times in the BL-3 laboratory while BL-3 experiments are in
progress. Open-toes shoes are not worn by any personnel entering the BL-3
laboratory. Additional personal protective equipment may be necessary based on
the particular activity.
45. Reusable laboratory clothing is decontaminated before being laundered.
Procedure: ___________________________________________________________
46. Face protection is worn for manipulations of infectious materials outside the
biosafety cabinet.
47. Respiratory protection is worn when aerosols cannot be contained such as outside
the biosafety cabinet. Enrollment in the University Respiratory Protection Program is
required.
48. Wearing personal protective equipment outside the BL-3 laboratory is prohibited.
49. Street outer-wear is not kept in the BL-3 laboratory.
50. The laboratory is kept neat and clean.
NO
Experimental Animals: Complete only if applicable.
YES
51. Laboratory animals held in the ABSL-3 Suite are housed in total containment.
The Vivarium Management, the Biosafety Officer, the BSL-3 Director, and the IBC
UCAR Coordinator must approve all other caging.
52. All laboratory personnel housing animals in the ABSL-3 Suite must be BSL-3
Authorized Users as per the BSL-3 Program.
53. All laboratory personnel housing animals in the ABSL-3 Suite must successfully
complete the ABSL-3 training program.
54. When required, approved respirators are worn in rooms containing experimental
animals. Enrollment in the University Respiratory Protection Program is required.
NO