Biological Safety in
the Laboratory
Robert J. Hashimoto, CBSP
The University of Minnesota
Minneapolis, Minnesota
November 15, 2007
Introduction
The purpose of this training session is to familiarize
you with the fundamentals of biosafety as it relates
to the Office, Vice President of Research and the
Department of Environmental Health and Safety.
We will discuss:
 Institutional Biosafety Committee (IBC) Review
 The elements that are reviewed on the application
form including but not limited to:
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Classification of Agents
Risk Assessment/Management
Work Practices/Engineering Controls/Personal Protective
Equipment
The Key Players
Biosafety Program Points of Contact
Office, Vice-President of Research
 Melissa Nellis: (612) 626-5892
 Julie M. H. Quinn: (612) 626-2161
DEHS
 Darlene Charboneau: (612) 624-5479
 XiaoHong Chen: (612) 624-5495
 Claire Kari: (612) 626-2145
IBC Forms

The reviews two key forms at a convened
meeting:
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The Recombinant DNA Form,
http://www.research.umn.edu/ibc/download/rDNA
%20Form.doc
The Infectious Agents Form,
http://www.research.umn.edu/ibc/download/Infecti
ous%20Agent%20Form.doc
IBC Forms

The submission of the application may require that
you write a scope of work and complete templates.
Other safety requirements:
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Section 6-10 on the Recombinant DNA Form
Section 6-8.5 on the Infectious Agents Form
The remainder of this presentation will discuss the
considerations that IBC will review in your
application and what you should include on the
safety sections of the forms.
Review the UMN Biosafety Manual for more
information

http://www.dehs.umn.edu/bio_pracprin.htm
Recombinant DNA Definition
Section I-B of the Guidelines

In the context of the NIH Guidelines,
recombinant DNA molecules are defined as
either:
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molecules that are constructed outside living cells
by joining natural or synthetic DNA segments to
DNA molecules that can replicate in a living cell,
or
molecules that result from the replication of those
described in (i) above.
Infectious Agent Definition
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Infectious or pathogenic agents capable of
causing disease in healthy humans, plants
and animals including but not limited to:
Bacteria
Virus
Fungi
Parasites
Rickettsia
Classification of Agents


The Centers for Disease Control and
Prevention (CDC) in the guideline,
Classification of Etiologic Agents on the
Basis of Hazard, classify agents on a scale
from Class 1 through Class 5.
However, the NIH Guidelines cites the World
Health Organization Risk Group
Classification from Risk Group 1 to Risk
Group 4, which will be discussed in the next
training session.
Risk Assessment

Once the investigator has decided on the agent or
recombinant molecule, then he or she must conduct
an assessment of risk. This assessment shall be
based on the following:
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Virulence/pathogenicity/infectious dose
Environmental stability
Route of spread, communicability
Quantity/concentration/volume used
Vaccine/Treatment availability
Allergenicity
Review Section IX of the UMN Biosafety Manual.
Risk Assessment
Routes of Exposure

The assessment of risk will include common routes
of exposure:
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Parenteral Inoculation (needle stick)
Surface Contact (contaminated work area)
Ingestion (food in lab)
Inhalation (aerosol generating procedures)
Mucous Membrane (aerosol droplets in face)
The IBC will review the agent usage and determine
the most likely routes of exposure (e.g. HIV,
bloodborne pathogen-percutaneous)
Public Health Agency of Canada MSDS

http://www.phac-aspc.gc.ca/msds-ftss/msds84e.html
Risk Assessment
Experimental Procedures

The IBC will also review the application for
the procedures performed to assess risk.
This includes but is not limited to the
following:
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Handling of sharps
Aerosol generating procedures
Volume of culture used
Transport of agents
Use of chemicals and radionuclides
Use of animals
Any open bench top work
Disposal of medical waste
Containment

The IBC will then evaluate containment of the
agents. What is containment?

The application of combinations of laboratory
practice and procedure, laboratory facilities, and
safety equipment when working with potentially
infectious microorganisms.
Biosafety Levels
Containment=biosafety levels.
The CDC guideline, Biosafety in Microbiological
and Biomedical Laboratories describe four
biosafety levels which consist of combinations of
laboratory practices, safety equipment and
laboratory facilities as well as the CDC hazard
classification of the agent and the type of
procedures used in the experiment. For
recombinant DNA experiments, refer to
Appendix G of the NIH Guidelines.
Biosafety Level 1

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Agent work conducted at Biosafety Level 1
(BL-1) involves little or no known potential
hazard to laboratory personnel and the
environment.
Work is generally conducted on the open
bench top and special containment
equipment, such as a biosafety cabinet, is not
required.
E-coli K-12 is an example of a BL-1
organism.
Biosafety Level 2
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Agent work conducted at Biosafety Level 2 requires
that biosafety cabinets are used whenever aerosol
generating procedures are performed.
Training and handwashing are also required.
Access to the laboratory is limited while the
experiment is in progress.
The Hepatitis B Virus is an example of a BL-2
organism.
Biosafety Level 3
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Agent work conducted at Biosafety Level 3 is
required for agents that may cause serious or lethal
disease by the inhalation route.
All BL-3 procedures are conducted within a
biosafety cabinet; the lab has special engineering
features to prevent a release of the BL-3 agent to
the environment. Lab workers may need to wear a
respirator.
Mycobacterium tuberculosis is an example of a BL-3
agent.
Biosafety Level 4
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Agent work conducted at Biosafety Level 4 is
necessary for all work with dangerous and
exotic agents which pose a high individual
risk of aerosol transmitted laboratory
infections and life threatening disease. Lassa
Fever Virus is a BL-4 agent.
There is currently no Biosafety Level 4
research conducted at the University of
Minnesota.
Minimization of Risk

Once exposure determination has been completed,
then ways to minimize risk are evaluated. This
includes but is not limited to the following:
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Work Practices/Engineering Controls
Personal Protective Equipment
Disinfection/Medical Waste Disposal
Medical Surveillance
Training (at the laboratory level)
Both the investigator and the IBC must review and
determine appropriate measures-it is the reason
why the application must be completed in detail.
Minimize Sharps Usage
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Needles must not be recapped, bent,
sheared or removed from a
disposable syringe.
All used sharps must be placed in a
rigid, hard-plastic, puncture-resistant
container for disposal.
Substitute plastic for glass whenever
possible.
Review Section XI of the UMN
Biosafety Manual
Handwashing
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Hands should be washed:
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immediately and thoroughly with a disinfectant
hand soap, if contaminated with biological agents.
after gloves are compromised or removed
with water after a puncture wound
If a sink is not available (e.g. equipment
room), consider the use of disinfectant
towelettes in these areas.
Minimize Aerosol Generation
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To avoid inhalation exposure to agents, workers
must minimize the potential generation of aerosols.
These procedures include but are not limited to:
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sonicating
centrifuging
grinding
blending, mixing
vortexing
Work on the open bench top should be limited
whenever conducting these procedures.
Review Section XIV-B of the UMN Biosafety Manual
Use Biosafety Cabinets
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Biological safety cabinets are:
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used for product/personnel protection
used for aerosol generating procedures
disinfected after use
certified annually
not recommended for chemical or radioisotope
usage.
Review Section III of the UMN Biosafety
Manual for more information.
Use Mechanical Pipettes
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Mechanical pipetting devices should be used
for manipulating all liquids in the laboratory.
Never mouth pipet.
Decontaminate Work Surfaces
Laboratory work surfaces should be
decontaminated with an appropriate chemical
disinfectant after a spill of biohazardous
materials and when work activities are
completed.
Disinfectant Selection
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Disinfectants must be selected on a case by case
basis to ensure efficiency.
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Quaternary Ammonia Compounds
Chloride Compounds (Bleach)
Iodophores (Wescodyne)
Phenolics (Amphyl))
Alcohols (70% Ethanol)
Formaldehyde/Glutaraldehyde
Review Section XVII of the UMN Biosafety Manual
for more information.
Infectious Waste Disposal
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Review the Infectious Waste Management
Plan in the UMN Biosafety Manual, Chapter
XVII-D
Ensure that:
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infectious waste is placed in a red bag lined bin.
sharps waste is placed in a sharps container
mixed chemical-biological waste is handled as
chemical waste
mixed radioactive-biological waste is handled as
radioactive waste
No Eating, Drinking, or Smoking
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Hand to mouth transmission of disease is a
common route of exposure while handling
biological agents.
Avoid eating, drinking, or smoking in
laboratory work areas.
Wash hands with disinfectant soap prior to
leaving the work area.
Do not store or heat/chill food or beverages in
the lab.
Personal Protective Equipment
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Use personal protective equipment to prevent
skin/mucous membrane exposure during agent
use, such as:
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Gloves
Safety glasses/face shield
Lab coat
Closed toe shoes/foot covers
Respiratory protection (BL-3 agents)
Review the UMN Biosafety Manual, Chapter XIIIE for more information
Hazard Communication
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Biohazard labels shall be placed on:
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Biohazard signs shall be placed on:
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the surface of all equipment (freezers, incubators,
refrigerators) which may be contaminated with
biohazardous materials.
sample transport outer containers.
medical waste bins
the outer door of BL 2 labs.
medical waste storage areas
Review Section XIII-B of the UMN Biosafety
Manual for more information.
Medical Surveillance
Requirements-Vaccinations
Vaccinations are available for some
organisms and should be offered when
feasible. However, some members of
the population may be at risk for medical
complications from the vaccination (e.g.,
excema-vaccinia vaccinations);
therefore it is extremely important that a
physician be consulted prior to the
administration of any vaccine.
Medical Surveillance
Some Available Vaccinations
Organism
Disease
Bordetella pertussis
Clostridium tetani
Corynebacteria diptheriae
HBV
Influenza
Mycobacterium tuberculosis
(Bacille Calmette Guerin or BCG)
Neisseria meningitidis
Polio Virus
Rubivirus
Salmonella typhi
Varicella Zoster
Variola major
Whooping Cough
tetanus
diptheria
hepatitis
Flu
-TB
meningococcal disease
Poliomyelitis
Rubella
Typhoid Fever
Chicken Pox
Smallpox
Biological Spills
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If a spill of biological material occurs:
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Always tend to injured person-Call 911
Notify supervisor and DEHS (626-6002)
Initiate clean up procedures
Notify the Office of Vice President of Research
as a report may need to be sent to NIH OBA.
Refer to the UMN Biosafety Manual, Chapter
I-C
Emergency Procedures
needlestick, surface exposure
 wash your skin with soap and water. Review Section
I-D of the UMN Biosafety Manual
splash in the eyes
 wash at an eyewash for 15 minutes.
Other injury
 Call 911
The IBC Application
Process
The Correct Form
Required Templates
Frequently Asked Questions
The IBC Application Process

To initiate the review process, University of
Minnesota researchers must submit an IBC
application form to the IBC Office. Obtain the
appropriate form based on your work:
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Recombinant DNA and gene transfer form
Infectious Agents Form
Biological Toxin Form
IBC approval is required PRIOR to the
initiation of experimental procedures.
The IBC Application Process
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In addition to the appropriate application
form, the following items must accompany
the completed form:
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Decontamination and Spill Cleanup Template
Biological Waste Disposal
Other Required Attachments Requested in the
Application or information requested by the IBC
upon review of the application form.
The IBC Application Process
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All submissions must be sent electronically to
ibc@umn.edu from the Principal Investigator's email account.
Projects are reviewed by the Institutional Biosafety
Committee. The IBC meets once a month to review
projects. The Committee may request modification
of procedures and may direct the Biosafety Officer
to inspect labs or other facilities.
An electronic approval letter will be sent to the
Principal Investigator when all requirements have
been met.
The IBC Application Process
Things to Remember

Q: How long will the IBC approval process take?
A: If an application is received on or before a
submission deadline, it will be reviewed at the next
IBC meeting, which falls approximately two weeks
after the deadline. The Principal Investigator will
receive notice of the meeting results approximately
7-10 business days after the meeting. Projects that
are assigned stipulations will take an additional
amount of time, depending on how quickly the
stipulations are met and when they can be
reviewed, either by the Biosafety Officer or by the
full committee.
The IBC Application Process
Things to Remember

Q: How should you respond to stipulations from
the IBC?
A: Responses to IBC stipulations should be
submitted electronically to ibc@umn.edu. Please
submit your response in the form of a letter,
addressing each point in the order in which they are
presented in the stipulations letter from the
IBC. Responses to stipulations should be attached
as a Microsoft Word document or PDF and sent
directly from the Principal Investigator’s University of
Minnesota e-mail account.
The IBC Application Process
Things to Remember

Q: Does research that is exempt from the NIH
Guidelines for Research Involving Recombinant
DNA Molecules require IBC review?
A: Yes, the University of Minnesota Board of
Regents Policy requires that University faculty, staff
and students using any potentially hazardous
biological agents (recombinant DNA, infectious
agents and/or biologically-derived toxins) in
research or teaching activities apply to the
IBC. Even if recombinant DNA activities are exempt
according to Section III-F of the NIH Guidelines, the
activities must be reviewed by the IBC.
The IBC Application Process
Things to Remember

Q: In what order should I submit my IACUC and
IBC applications?
A: There is no prescribed requirement for the order
in which IACUC and IBC application are submitted;
however, IACUC final approval will not be granted
until IBC approval has been received. It is advisable
to submit your IBC proposal either before or at the
same time as when you submit your IACUC
proposal to avoid delays. You need a separate form
submitted to each committee as each committee
reviews different issues.
Accident Reporting
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Accident reporting is critical for several reasons:
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Risk management
Reporting to local regulatory agencies (e.g. Department of
Public Health)
IBC NIH obligations whenever recombinant DNA molecules
are involved
Serious adverse events-human subjects
The Principal Investigator must report accidents
involving biological agents and recombinant DNA to
the IBC and DEHS within 24 hours. The Office of
Vice President for Research and DEHS will
determine who may need to be notified.
Conclusion
The University of Minnesota Department of
Environmental Health and Safety and the
Office of the Vice President of Research
jointly participate in the review of biological
research. If you must perform experiments
with recombinant DNA or infectious agents,
please remember the following:
Conclusion
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Go on-line to download the appropriate forms
and safety templates;
Allow sufficient time for the appropriate
institutional committees to review your work;
Ensure that the form has adequate
information for the IBC to assess your workinclude the safety considerations and
assessment of risk pertinent to the agent or
recombinant molecule that you are using.
Conclusion
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If you have any questions on the
administrative process of IBC approval,
please contact Melissa Nellis (626-5892)
If you have any questions on biosafety
containment or DEHS related safety issues,
please contact Darlene Charboneau (6245479).