Standard Operating Procedure (SOP) for Staphylococcal Enterotoxin
University of California, Irvine
Purpose
This SOP describes the techniques and procedures for safe and proper handling, storage
and preparation for experimental use, and disposal of Staphylococcal Enterotoxin.
Receiving
1. When the shipment is received the specification sheet goes in the Select Agents
Notebook with a date on the log.
2. Staphylococcal Enterotoxin is placed in a locked refrigerator in the lab until aliquoted
for use.
Minimum Personal Protective Equipment (PPE)
Non-permeable nitrile or latex gloves
Disposable long-sleeved lab coat
Safety glasses or goggles
When handling toxins that are percutaneous hazards (irritants, necrotic to tissue, or
extremely toxic by dermal exposure), select gloves that are known to be impervious to
the toxin. Consider both the toxin and the diluent when selecting gloves or other
protective clothing. If infectious agents and toxins are used in an experimental system,
consider both when selecting protective clothing and equipment. Respiratory protection
may be required if aerosols may be generated and it is not possible to use containment
equipment or other engineering controls.
Placarding
When toxins are in use, the room should be posted to indicate “Toxins in Use-Authorized
Personnel Only”. Any special entry requirements should be posted on the entrance(s) to
the room. Only personnel whose presence is required should be permitted in the room
while toxins are in use.
Procedures for Handling of Staphylococcal Enterotoxin
Inspect the containers for damage during shipment. If damaged contact the vendor
immediately, and decontaminate and dispose of the shipment immediately. If the vials
were not damaged excess packing materials may be disposed in the regular trash. Log in
the select agent shipment on the select agent inventory sheet. All packages containing
select agents should be opened in a fume hood, glove box or ducted biosafety cabinet.
Notify the sender within 24 hours of receipt of the toxin. Maintain a copy of the
completed EA-101 form for lab records and fax copy to the Biosafety Officer at (949)
824-3383.
All high-risk operations should be conducted with two knowledgeable individuals
present. Each must be familiar with the applicable procedures, maintain visual contact
with the other, and be ready to assist in the event of an accident.
If the Staphylococcal Enterotoxin is received as a powder, place a pad on the inside of the
fume hood, glove box or ducted biosafety cabinet in order to minimize the spread of
contamination during the weighing process. Ensure the fume hood, glove box, or ducted
Biosafety cabinet is working properly with an inward airflow prior to initiating work. All
work should be performed within the operationally effective zone of the fume hood or
biosafety cabinet. When handling dry forms of the powder that are electrostatic, do not
wear gloves (such as latex) that generate static electricity. Use a glove bag within a fume
hood or biological safety cabinet, a glove box or class III biosafety cabinet.
If you are aliquoting samples, place an absorbent pad on the bottom of the fume hood or
ducted biosafety cabinet in order to contain any potential spills. Ensure the fume hood,
glove box, or ducted Biosafety cabinet is working properly with an inward airflow prior
to initiating work. All work should be performed within the operationally effective zone
of the fume hood or biosafety cabinet. Resuspend the toxin by extremely careful and
slow titration, rinsing down the walls of the tube in the process. Avoiding foaming and
aerosolization. Aliquot 5 mls of 10% bleach solution into a 50 ml conical tube and place
the open tube in the tube rack. This tube will serve as a waste receptacle for
contaminated pipet tips. When vacuum lines are used with systems containing toxins,
they should be protected by a HEPA filter to prevent entry of the toxin into the lines.
Sink drains should be similarly protected when water aspirators are used.
Place the toxin vials in an unbreakable, clean secondary storage container and transfer to
storage at the appropriate temperature. Decontaminate the exterior surfaces of all
materials leaving the biosafety cabinet, including the closed aliquot tubes with a 10%
bleach solution. Decontaminate all work surfaces with a 10% bleach solution. Until the
fume hood, glove box or ducted biosafety cabinet is decontaminated, the equipment
should be posted to indicate that toxins are in use, and access to the equipment and
apparatus is restricted to authorized personnel. Toxins should be transported only in
leak-proof secondary containers.
Standard sharps must not be used with Staphylococcal Enterotoxin unless specifically
approved by Environmental Health and Safety. If it is absolutely necessary to use sharps
with Staphylococcal Enterotoxin, sharps with engineering controls must be used.
Storage of Stock
1. Have EH&S pickup empty bottle for disposal.
2. When all of the aliquots are gone then log into Select Agent log that all aliquots have
been used.
Engineering Controls
Engineering controls such as fume hoods and biological safety cabinets must be used as
primary containment in order to limit personnel exposure to Select Agents. Engineering
controls must be inspected in order to ensure efficient removal of hazard, and must have a
visual indication of airflow, and alarms to indicate that airflow has fallen below
acceptable standards. Notification of laboratory personnel must be made prior to any
maintenance that will impact the capture velocity of ventilation systems. Local exhaust
ventilation systems including laboratory-type chemical fume hoods and biosafety
cabinets must be certified upon installation, after maintenance and annually thereafter.
Administrative Controls
Written Standard operating procedures should be developed for all laboratories
possessing, using, transferring or receiving select agents in order to ensure consistently
safe work practices are being used. Standard Operating Procedures must be reviewed
annually or as necessary to reflect changes to procedures. Standard Laboratory Practices
for Guidelines for Work with Toxins of Biological Origin are addressed in the BMBL
Appendix I. http://www.cdc.gov/od/ohs/biosfty/bmbl4/b4ai.htm
Security
Select Agents must be stored in a locked facility within a locked freezer, refrigerator, or
cabinet when not in use. An inventory control system should be in place. Access to
areas containing toxins should be restricted to those whose work assignments require
access. Principal Investigators are responsible for ensuring that all laboratory workers
and visitors understand security requirements and are trained and equipped to follow
established procedures.
Laboratory and animal areas with select agents must be locked at all times. All entries
(including entries by visitors, maintenance workers, and others needing one-time or
occasional entry) should be recorded by signature in a log book. Only workers required
to perform a job should be allowed in laboratory areas, and workers should be allowed
only in areas and at hours required to perform their particular job. Access for students,
visiting scientists, etc., should be limited to hours when regular employees are present.
Access for routine cleaning, maintenance, and repairs should be limited to hours when
regular employees are present. The laboratory must be cleared of hazardous materials by
EH&S prior to maintenance work or cleaning.
Freezers, refrigerators, cabinets, and other containers where stocks of select agents are
stored must be locked when they are not in direct view of the workers (e.g., when located
in unattended storage areas).
Decontamination and Disposal
All liquid waste must be decontaminated using a 10% bleach solution for a 30-minute
contact time and may be drain disposed with plenty of water. Dispose decontaminated
liquid waste down the drain with plenty of water. Dry waste must be placed in double
red biohazard bags autoclaved in an autoclave approved for Medical Waste treatment at
121 C for 60 minutes at 18 psi. Any destruction of Select Agents must be witnessed by
the RFO. Contaminated and potentially contaminated protective clothing and equipment
should be decontaminated using methods known to be effective against the toxin before
removal from the laboratory for disposal, cleaning or repair. If decontamination is not
possible/practical, materials (e.g. used gloves) should be disposed of as hazardous waste.
Materials contaminated with infectious agents as well as toxins should be also be
autoclaved or rendered non-infectious before leaving the laboratory.
Spill Clean up
If a spill occurs outside of the fume hood or ducted biosafety cabinet, notify lab personnel
and evacuate immediately. Close the doors as you exit to allow the aerosols to settle.
Notify EH&S immediately! Do not re-enter the facility.
Personnel decontaminated the spill must wear the minimum PPE of safety goggles,
disposable lab coat, double gloves and shoe coverings. Respiratory protection may be
necessary based on a risk assessment of the Select Agent.
Cover the spill area with absorbent material and cover with 10% bleach solution for a 30minute contact time. Do not splash the material in your face, and attempt to limit
exposure. Clean up the material and place in double red biohazard bags. Ventilate the
area and wash the spill site again after decontamination is complete. Notify your
supervisor and EH&S of the spill immediately at 824-5073.
Medical Surveillance
Medical surveillance may be required for laboratory personnel based on potential risks of
exposure. The PI should determine the need for medical surveillance with the assistance
of Environmental Health and Safety.
Post-Exposure Management
In the event of an exposure to Staphylococcal Enterotoxin Immediately call EH&S at
824-5073 or 911 from a Campus Phone. After hours (before 8AM or after 4:30pm) call
(714) 456-7890 and request the Infectious Disease Fellow On-Call or go to the nearest
hospital emergency room.
Symptoms of Staphylococcal Enterotoxin Exposure include:
 Fever
 Chills
 Headache
 Myalgia
 Non-Productive Cough
 Shortness of Breath
 Nausea
 Diarrhea
 Vomiting
 Septic Shock
Treatment:
 No specific vaccine is available
 Supportive care
 Anticipation of the ventilation of respiratory system for acute cases
Training
All personnel possessing, using, transferring, or receiving select agents must have
training as to the symptoms of toxin exposure, Post-Exposure Management Protocol, spill
cleanup and decontamination, proper use of engineering, administrative and work
practice controls and personal protective equipment, and security requirements for select
agent possession and use. Personnel must be trained on the Chemical Hygiene Plan and
the Select Agent specific Standard Operating Procedures. Training is required before
initiation of research involving Select Agents and annually thereafter.
For Additional Information
Please contact the Biosafety Officer, Ms. Susan Weekly at 824-9888 or sweekly@uci.edu
for additional information or assistance.