Summary of Guidelines for the Safe Use of the Flow Cytometry Facility
The Flow Cytometry Facility operates as a Biosafety Level 2 laboratory. These guidelines are reviewed and revised if necessary each year and each facility user is responsible to review them on a regular basis to insure they are in compliance. Occasionally, some cell sorting experiments may operate in the cell sorter room under BSL-2+ conditions. This will be determined by the Flow Core Staff.
1) Admission to the laboratory is restricted to authorized personnel when work with human samples or a known infectious agent is in progress.
2) All cells must be assigned appropriate BSL levels for research use by the Biological Safety and Recombinant DNA Committee (Ralph Keil, Ph.D., Chair) before they are allowed to be used in the Flow Cytometry Core. Additional information can be found on the Safety Infonet website.
3) No BSL2+ samples are allowed to be run on the analyzers.
4) Before any cell sorting experiment can be scheduled, the Cell Sorting Experiment Form must be completed, signed and presented to the operator. This allows time for appropriate evaluation of biohazards under high speed sorting conditions. This must be done at least a week before the proposed experiment.
5) Some unfixed, primary human and primate cells and other BSL2 cells to be used on the FACSAria High Speed Cell Sorter may be designated as BSL2+ by the Flow Biosafety Committee. In these cases, strict adherence to the BSL2+ SOP will be required (located in the Flow Core). These same cells MAY be used under BSL2 conditions on the Cell Sorter if they are fixed according to accepted methods of inactivating all biohazards. (See note on Special Requirement for Sorting Primary Human Cells below.)
6) Whenever possible, samples of human or primate origin to be run on any cytometer should be fixed according to a process known and documented to inactivate HIV and other biohazards.
7) All procedures must be performed carefully to minimize the creation of aerosols.
8) All samples and time used on the instruments must be logged into the facility in Proteus. Samples that have been fixed for analyzing or sorting on the Cell Sorter may be monitored and recorded to insure that the fixation is adequate.
9) Work surfaces are to be decontaminated with 10% household bleach (FACS Clean) after any spillage of sample or hazardous material and after each user completes his/her work.
10) Instrument decontamination shall be performed according to protocols posted on each instrument.
11) All spills, however small, must be recorded in the accident book and reported to the Lab Manager.
12) All liquid and solid waste and disposal is to be removed from the laboratory as biohazardous material by each user. This includes all nuclear staining reagents such as PI, EB, AO, etc.
13) Gloves must be worn whenever handling or running biohazardous samples on the cytometer.
14) Hands must be washed after handling biohazardous materials and before leaving the laboratory.
15) Laboratory coats must be worn in the work area. These coats should be removed before leaving the laboratory.
16) Neither syringes nor hypodermic needles should be used in the facility without the consent of the Laboratory Manager. Glass objects should only be used when absolutely necessary.
17) No eating, drinking, smoking or applying cosmetics in the work area.
18) No mouth pipetting.
19) No animals are permitted within the laboratory.
20) All biohazard samples must be transported to and from the Flow Core in double sealed containers per College of Medicine safety policy.
In addition to the above guidelines and because this facility uses Class 3a through Class 4 lasers, the following Laser Safety guidelines shall also be observed:
- Access to the laboratory will be limited to only those persons necessary to the running of experiments when any laser is operating in the UV mode or when it is necessary to operate a laser without protective shielding.
- In such cases, the appropriate protective goggles or glasses will be made available and must be worn by those in the work area.
- Only qualified personnel shall operate or adjust any laser.
In the event of a potential retrovirus exposure
This item refers to exposure of skin or mucous membranes to infectious materials, and is modeled after the NIH program, "3 Emergency Steps to Take in the Event of a Potential HIV Exposure". Remember that the risk of infection is very dependent upon the titer of virus and the route of exposure. Thus the risk of infection by contact of intact skin with infectious body fluids is probably truly zero, and although the risk of infection by contact of mucous membranes or non-intact skin with infectious body fluids may be extremely small, it is likely to be no-zero.
The 3 emergency steps are:
- Immediately initiate first aid at the work site.
- Contaminated skin should be meticulously cleaned for 10 minutes using a povidone iodine solution (such as Betadine) and copious amounts of water.
- Contaminated eyes and mucous membranes should be irrigated for 5 minutes using water.
- Notify your supervisor, if he or she is immediately available. More importantly, go on to step 3.
- Report to the hospital Emergency Room to activate the invasive accident protocol (needlestick protocol), which will include evaluation, counseling, and provision of antiretroviral treatment if deemed appropriate. Do this immediately (i.e. within the half-hour).