Rules for Working with Microorganisms

Precautionary Proceedures

• Do not eat, drink, smoke, apply cosmetics or lip balm, or store food in the laboratory.
• Bring any necessary equipment, laboratory instructions and your lab notebook to the work bench. A flow chart and/or notes may help you carry out the lab work with a minimum of page shuffling.
• Wear a protective laboratory coat or apron when you are working with cultures, and avoid wearing long, full sleeves if possible.
• Tie long hair back or put it up. If hair hangs loose, it becomes a contamination hazard and also may catch on fire in the bunsen burner flame.
• Carry and store cultures of microorganisms in racks or baskets. Do not leave cultures on the table or in unmarked areas when the laboratory session is completed.
• Place cultures to be discarded in racks or trays designated for contaminated material; these racks should be clearly labeled. All such materials should be autoclaved before further handling, discarding, or washing.
• Decontaminate work surface after spills and at the beginning and end of each laboratory period with the disinfectant provided in the laboratory.
• Cover small spills with paper towels and soak the towels well with disinfectant. Let the towels stand for one-half an hour. Place these materials in a container with disinfectant or in a plastic bag which can be sealed. Autoclave container before discarding.
• Mix liquid cultures gently to avoid foaming and splashing which may produce an aerosol. Do not mix cultures by bubbling expiratory air through the liquid with a pipette.
• Never pipette cultures by mouth; mechanical pipetting devices are to be used. Place contaminated pipettes in a pan which contains an appropriate disinfectant.
• Wash hands carefully with soap after any possible contamination and before leaving laboratory. Dry hands thoroughly after washing.
• Open cuts should be covered with bandages; if they are on the hands, wear disposable gloves.
• Individuals who have special health problems such as diabetes or suppressed immunity from disease or therapy should be encouraged to discuss them privately with the instructor.
• Develop the habit of keeping your hands away for your mouth, nose, eyes, and face to prevent self-inoculation.
• Before centrifuging, inspect tubes for cracks, Whenever possible, use autoclavable plastic centrifuge tubes with screw caps. Avoid filling the tube to the point that the rim becomes wet with the culture.
• Avoid the use of hypodermic syringes. Use a pipette whenever possible. If a syringe is needed, use a needle-locking hypodermic syringe.
• Before and after injecting an animal, swab the site of injection with disinfectant.
• Shake broth cultures in a manner that avoids wetting the plug or cap.
• Periodically clean out deep freeze and dry-ice chests in which cultures are stored to remove broken ampules and tubes. Use rubber gloves and respiratory protection during the cleaning.
• Handling of materials infectious to man and other animals in the laboratory require the September 16, 2011 federal, state and local government.

Emergency Procedure For Working With Microorganisms:

• Spills
  In the case of spill, flood area with disinfectant provided in the laboratory. Dispose of paper towels with contaminated waste. Wipe counter dry with paper towels.
  For spills on carpeting, after flooding with disinfectant, clean up as directed by the instructor.
• Ingestion
  Do not swallow.
  Empty contents of mouth into sink.
  Rinse mouth with copious amount of water. 
  Report all ingestion to instructor.
• Exposure
  Eyes: Immediately WALK to eye wash station. Turn on water. Hold eyelids open and wash for at least two minutes. Inform instructor. 
  Cuts: Wash cut with water. Follow with disinfectant. Place a pad or clean material over the cut. Apply pressure directly over cut. Do not put mouth over the cut. Do not breathe on the cut.
  Needle injection: Report immediately to the instructor. The instructor will call infirmary and the infirmary will then determine the appropriate course of action. 

Biosafety Level 2 

Biosafety Level 2 is suitable for experiments involving agents of moderate potential hazard to personnel and the environment. It differs in that laboratory personnel have specific training in handling pathogenic agents and are supervised by competent scientists. Access to the laboratory is limited when experiments are being conducted, and procedures involving large volumes or high concentrations of agents or in which aerosols are likely to be created are conducted in biological safety cabinets or other physical containment equipment. 

Special Practices 

Contaminated materials that are to be decontaminated at a site away from the laboratory are placed in a durable leak-proof container which is closed before being removed from the laboratory.

• Access to the laboratory is limited by the laboratory supervisor when experiments are being conducted. In general, persons who are at increased risk of acquiring infection or for whom infection may be unusually hazardous are not allowed in the laboratory or animal rooms. Persons at increased risk may include children, pregnant women, and individuals who are immunodeficient or immunosuppressed. The supervisor has the final responsibility for assessing each individual circumstance and determining who may enter or work in the laboratory.
• The laboratory supervisor will assure that only persons who have been advised of the potential hazard and meet any specific entry requirements (e.g., immunization) may enter the laboratory or animal rooms.
• When infectious materials or infected animals are present in the laboratory or animal rooms, a hazard warning sign, incorporating the universal biohazard symbol, is posted on all laboratory and animal room access doors and on such other items (i.e., equipment, containers, materials) as appropriate to indicate the presence of viable infectious agents. The hazard warning sign should identify the agent, list the name of the laboratory supervisor or other responsible person(s), and indicate any special requirements for entering the area (immunization, respirators, etc.).
   

Biohazard Warning Symbol:

An insect and rodent control program is in effect.

Laboratory gowns, coats, or uniforms must be worn in the laboratory but must not be worn to a lunchroom or outside the building in which the laboratory is located.

• Animals not involved in the experiment being performed are not permitted in the laboratory.
• All wastes from laboratories and animal rooms must be appropriately decontaminated before being disposed.
• The use of hypodermic needles and syringes is restricted to lavage, parenteral injection and aspiration of fluids from laboratory animals and diaphragm vaccine bottles. Hypodermic needles and syringes are not used as a substitute for automatic pipetting devices in the manipulation of infectious fluids. Serial dilutions of infectious agents should not be done in diaphragm bottles with needles and syringes because of the hazards of autoinoculation and of aerosol exposure. Cannulas should be used instead of sharp needles whenever possible.
• If activities of lesser biohazard potential are conducted in the laboratory concurrently with activities requiring Biosafety Level 2, all activities will be conducted at Biosafety Level 2.
• Gloves should be worn for all procedures requiring the handling of infectious materials or infected animals. Holding small laboratory mammals with forceps when they are receiving injections or otherwise being handled provides an additional level of protection for personnel.
• Serological procedures with inactivated antigens shown to be free of residual infectivity can be performed on the open bench.
• All spills, accidents, and overt or potential exposures to infectious materials must be immediately reported to the laboratory supervisor. A written record must be prepared and maintained. Appropriate medical evaluation, surveillance, and treatment
• Baseline serum samples should be collected from and stored for all laboratory and other at-risk personnel. Additional serum specimens may be collected periodically depending on the agents handled or the functions of the facility.
• A safety or operations manual which identifies known and potential hazards and which specifies practices and procedures to minimize or eliminate such risks should be prepared or adopted. Personnel should be advised of special hazards and are required to read and follow standard practice and procedures.
   

Containment Equipment

Biological safety cabinets (Class I, II, or III) or other appropriate personnel protective or physical containment devices are used whenever:

• Procedures with a high potential for creating aerosols are conducted. These may include centrifugation, grinding, blending, vigorous shaking or mixing, sonic disruption, opening containers of infectious materials whose internal pressures may be different from ambient, intranasal inoculation of animals, and harvesting infected tissues from animals or eggs.
• High concentrations or large volumes of infectious agents are used. Such materials may be centrifuged in the open laboratory if sealed heads or centrifuge safety cups are used and if they are opened only in a biological safety cabinet.