Lentiviruses - University of Southern California

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Dec 14, 2013 (3 years and 9 months ago)

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Exposure Control Plan for
:
Lentiviral Vectors


Containment Level: BSL
2


I. HAZARDS

Lentiviruses are a subset of retroviruses
with the ability to integrate into host chromosomes and to infect
proliferating and non
-
proliferating cells.
Thes
e viruses can cause severe immunologic and neurologic disease in their natural hosts.
Most of the lentiviral
vectors presently in use are derived from the Human Immunodeficiency Virus (
HIV
).

The major risks to be considered for research with HIV based lent
iviral vectors are the potential for generation of replication
-
competent
lentivirus (RCL) and the potential for oncogenesis. These risks can be mitigated by the nature of the vector system and its
safety
features (i.e.,
use of multiple plasmids for vector

packaging, deletion of overlapping sequences

between the packaging plasmids,
minimalist design of the vector backbone
)

or exacerbated by the nature of the transgene insert (i.e., oncogene) encoded by the vector.

Although most viral vectors used for gene d
elivery are not replication competent, virions or infectious particles produced with these
agents are
capable of infecting lab personnel
. Consequently, these materials should be treated and handled with appropriate care



strict adherence to BSL2 containm
ent is a necessity
.



II. MODES OF TRANSMISSION

The most likely modes of transmission
within the lab
include penetration of the skin via
puncture

or absorption through scratches, cuts,
etc., and direct contact with mucous membranes (eyes, nose and mouth)
.



III. ENGINEERING CONTROLS



A Certified Biosafety Cabinet

must be used
for
all manipulations

of the viral vector (i.e., pipetting, harvesting, infecting cells,
filling tubes/containers, opening sealed centrifuge canisters, shaking, mixing, etc.) and fo
r handling infected cells.



Safety Engineered Sharps
shall be used for
Lentivirus

injections
. In addition, the use of other sharps (i.e., Pasteur pipettes)
must be

eliminated

wherever possible.



For animal injections, the animal must be
restrained or anes
thetized



Biohazard
Sharps
C
ontainers

shall be available to dispose of sharps waste, including broken glass, needles, blades, etc.



When centrifuging, use aerosol containment devices such as
sealed canisters

that fit in the centrifuge bucket, covers for the
centrifuge bucket, heat
sealed tubes
, or
sealed centrifuge rotors
. Rotors should be removed and opened inside a BSC.
Centrifuge tubes should be filled and opened in BSC




An in
-
line HEPA filter

must be used for vacuum aspiration of spent media



IV. ADIM
INISTRATIVE/WORK PRACTICE CONTROLS



Access to the lab shall be restricted while work is in progress



A sign incorporating the universal biohazard symbol shall be posted at the entrance of the laboratory or tissue culture room
where
lenti
virus is used

(see l
ast page)



All lab personnel must be informed of the hazards of
Lenti
viruses



All lab personnel must be trained in proper handling, use, and disposal prior to working
Lenti
viruses



All lab personnel are advised to avoid rubbing eyes as a precautionary measure

against eye infections



All lab personnel will remove lab

coat, discard gloves, and wash hands before exiting the lab



V. PERSONAL PROTECTIVE EQUIPMENT



Lab coat

shall be worn while working in the lab



Safety glasses or goggles
shall be worn when handlin
g

Lenti
virus



Gloves

shall be worn while working
in
the lab



R
espirators

are required for
aerosol
-
producing procedures
performed outside of a biosafety cabinet
. Contact the EH&S
office at (323)442
-
2200 for fit
-
testing prior to use of respirators.





Revised
6/2/09

V
I
. D
ISINFECTION


General surface disinfection with
freshly prepared
10% bleach (1:10 dilution household bleach, such as clorox) allowing a contact time
of 15 minutes. For stainless steel surfaces, follow bleach disinfection with 70% ethanol wip
edown to avoid corrosion.

Liquid waste may be treated by exposing to bleach (final volume 10%) for 15 minutes before disposing into sink.



VII. DISPOSAL

Solid biohazardous waste, such as culture vials, plates, plastic tubes, etc., are disp
osed of into biohazard waste bins lined with red
bags for pickup by EH&S. Biohazard bins are labeled with biohazard stickers, and lids are to remain secured when not in use.

Sharps waste, such as broken glass, pasteur pipets, razor blades, and needles, ar
e disposed of into biohazard sharps containers.
When the fill line on the sharps container is reached, the lid will be secured and the container placed next to the biohazard

bin for
pickup by EH&S. To request a pickup, call the EH&S waste disposal line a
t (323)442
-
2225.



VIII. ACCIDENTAL SPILL

In case of spill:



Immediately notify others around you



Don appropriate personal protective equipment, such as gloves, labcoat, and safety glasses



Use forceps to remove any broken glass or other shar
p items, which should be placed into biohazard sharps containers



Cover the spill with paper towels or other absorbent materials



Apply 10% bleach directly onto and around the paper towels covering the spill



Allow 15 minute contact time before cleaning, star
ting at perimeter and working inwards towards center



Dispose of materials into biohazard bins



For large spills, you may contact the EH&S office for assistance at (323)442
-
2200



IX. EXPSOURE RESPONSE

In the event of an exposure, take the f
ollowing precautions:



Remove any contaminated clothing



Wash all affected areas; for eye exposures, rinse for 15 minutes in eyewash or flush area with water, for needle
-
stick or other
sharps exposure, wash wound area with soap and water for 15 minutes



Repor
t the exposure to your supervisor immediately



For students, seek treatment at the Eric Cohen Student Health Center:

H
ealthcare Consultation Center (HCC
)

1510 San Pablo St., Suite 104

(323)442
-
5631



For employees, seek treatment at Internal Medicine:




Healthcare Consultation Center II (HCC II)



1520 San Pablo St.



(323)442
-
5100



For any emergency, you may also contact the Department of Public Safety at (323) 442
-
1000



Healthcare personnel treating exposed patients should be inf
ormed of the nature of the agent (i.e.
,
lenti
viral vector

derived
from
HIV
-
1
)



Information on workers’ compensation can be found at this website:
http://capsnet.usc.edu/WC/
, or you may call: (213)740
-
6205



Incident
s should be reported to the Institutional Biosafety Committee as soon as possible:
IBC@caps.usc.edu

or (323)442
-
2200







Revised
6/2/09

X. SPECIAL PRACTICES FOR ANIMAL INJECTIONS




Laboratory staff must make arran
gements for housing these animals and for disposing of the contaminated bedding by
contacting the appropriate animal caretaker supervisor before dosing begins



Facility:
When animals are infected with
lentiviral
vectors, the Animal Biosafety Level of the p
roject will generally be
assigned to ABSL
-
2. This requires Biosafety Level
-
2 practice and facilities for procedures involving
lenti
viral vectors.



Signage:
Attached door sign must be posted on the door leading into the housing or procedure room. Cages mu
st be
labeled with the biohazard
cage card label with agent identification and injection date
upon injection of
agent
.

Signage/labels must remain in place for a minimum of 1 week after the date of injection/exposure.



Animal Excretion:
Infected animals my

excrete
lentivirus
.

Precaution must be taken not to create aerosols when emptying
animal waste material and when washing down cages, or cleaning the room with pressure hoses. Surfaces that may be
contaminated will be decontaminated ASAP with bleach solu
tion. This practice must be followed for one full week after
infection.



Cage Change:
Use a certified Class II biosafety cabinet when moving animal from dirty to clean cages. If multiple cages
are being changed in the same biosafety cabinet, cages with a
nimals infected with the
lentiviral
vectors should be changed
last. Spray the inside surfaces of each cage with bleach solution and leave the cages in the biosafety cabinet for 15
minutes before returning them to the cage washing facility. Decontaminate
the biosafety cabinet immediate after removing
the contaminated cages with bleach solution.



Bedding:

A biosafety cabinet or negative airflow cage changing station should be used when disposing bedding into
biohazard red bag. The red biohazard bag must be

sealed and placed i
nto the biohazard container.



Animal Carcass:

Infected carcasses should be placed in red biohazard bag and stored in designated freezer for EH&S
pick
-
up. EH&S must be informed when carcasses are put into freezer.
Revised
6/2/09


University of Southern

California

Laboratory Safety (323) 442
-
2200

ANIMAL HANDLER PRECUATIONS

Building:






Room Number:














The animals in this program are part of an approved experiment involving radioactive materials, biohazardous or
ganisms,
and/or chemical carcinogens. The
requirements

below apply to this experiment.


Principal Investigator:









Department:









Phone:







Emergency Contact Person:









Phone:










IACUC Protocol:






Identification of Hazard(s):
LENTI
VIRAL VECTOR

Animal Biosafety Level: 2


Required PPE

1.

Lab coat/coveralls

2.

Head cover

3.

Shoe covers

4.

Surgical Mask

5.

Safety Glasses (if animals handled ou
tside of a biosafety cabinet or ventilated cage
-
changing station)


Special

Instructions

1. All animals must be housed in micro
-
isolation cages.

2. Individual cages must be labeled with biohazard stickers upon injection of agent.

3. All animal handling m
ust occur inside certified biological safety cabinet.

4. All animal injections must occur in certified biosafety cabinet in a procedure room.

5
. Biosafety cabinet must be decontaminated with
5% bleach

after use or cage change.

6
. Contaminated bedding mu
st be disposed
into Biohazard Waste inside biosafety cabinet or negative
airflow cage changing station
.


7
.
Contaminated a
nimal carcasses must be disposed
in

a red biohazard bag
and place in the

designated
freezer. Inform EH&S
(HSC: x22225, UPC: x07215)

immediately for pick
-
up.

8
. Hands must be washed with soap and water upon leaving room.

Revised
6/2/09


AUTHORIZED PERSONNEL ONLY

WHEN LENTIVIRUS WORK IN PROGRESS


University of Southern California

Laboratory Safety (323) 442
-
2200

BIOSAFETY PRECUATIONS

Building:






Room Number:














Identification of Hazard(s):
LENTIVIRA
L VECTOR

Biosafety Level: 2



Principal Investigator:









Department:









Phone:







Emergen
cy Contact Person:








Phone:










IBC Protocol#:







Date:







Authorized Personnel: