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Biosafety
- 1. BIOSAFETY POOJA VERMA MSB/14/120 M.SC BIOTECH 2ND SEMESTER
- 2. BIOSAFETY: Preventing lab-acquired infections Bacteria Viruses Fungi Human blood, unfixed tissue Human cell lines Recombinant DNA
- 3. Guidelines NIH Guidelines for Experiments Involving Recombinant DNA Molecules Large Scale > 10 liters Biosafety in Microbiological and Biomedical Laboratories-NIH/CDC UM Biosafety Manual On DES webpage www.inform.umd.edu/des
- 4. Regulations OSHA Bloodborne Pathogens Standard Maryland Waste Regulations Shipping and packaging infectious substances DOT, UN, CDC, IATA
- 5. Levels of Containment BL1 - microorganisms that don’t consistently cause disease in healthy adults E. coli K12, S. cerevisiae, polyomavirus Basic laboratory Standard Microbiological Practices
- 6. Levels of Containment BL2 - microorganisms of moderate potential hazard, transmitted by contact, ingestion, puncture Salmonella, herpesvirus, human blood Basic laboratory Standard Practices PLUS
- 7. Levels of Containment BL2 - Standard Microbiological Practices Plus: Training in handling pathogens Access to lab limited Extreme sharps precautions Use of BSC for aerosols
- 8. Biosafety Cabinets Courtesy of the Baker Company
- 9. Use of Biosafety Cabinet Turn on fan 15 min before starting Don’t block grille Disinfect work surface With 70% etohnal Discard pipets inside cabinet Minimize movement of hands Avoid use of flame unless necessary Have cabinet certified annually
- 10. Clean Bench The clean bench provides product protection by ensuring that the work in the bench is exposed only to HEPA-filtered air. The clean bench is recommended for work with non- hazardous materials where clean, particle-free air quality is required. It does not provide protection to personnel or to the ambient environment.
- 11. Levels of Containment BL3 - microorganisms that cause serious disease, transmitted by inhalation M. tuberculosis, yellow fever virus, hantavirus, Y. pestis (plague) Containment lab: double door entry; directional airflow; all work in biosafety cabinet
- 12. Levels of Containment BL4 - microorganisms that cause lethal disease, with no known treatment or vaccine Ebola virus, Marburg virus Maximum containment lab; positive pressure ventilated suits (moon suits)
- 13. OSHA(occupational safety and health administration) Bloodborne Pathogens Standard Human blood, unfixed tissue, primary human cell culture, other potentially infectious materials HIV,HBV, HCV
- 14. OSHA Standard requires: Annual training Web-based program/DES homepage Free HBV vaccine Use of Universal Precautions
- 15. Universal Precautions Treat ALL human blood and unfixed tissue as if it contains HIV and HBV
- 16. Routes of Occupational Transmission Puncture or cut (needlestick, contaminated broken glass) Contact with broken skin Splash to mucous membranes of eyes, nose, mouth
- 17. Precautions for First Aid Wear gloves If conscious, have patient put pressure on wound Use one-way valve for CPR
- 18. Standard Microbiological Practices NOT permitted in laboratories: Eating Drinking Smoking Handling contact lenses Pipetting by mouth Storing food and drink
- 19. Standard Microbiological Practices ALWAYS wash hands: After handling microorganisms and animals After removing gloves Before leaving laboratory
- 20. Standard Microbiological Practices Discard needles, razor blades, and scalpel blades into red, puncture resistant sharps containers Dispose of broken glass into “broken glass” containers, never regular trash
- 21. Standard Microbiological Practices NEVER recap, bend, or break needles discard needles or sharps into biological waste bags discard needles into regular trash
- 22. Standard Microbiological Practices Decontaminate all biological waste (including BL1) before disposal Solid waste (Petri dishes, cultures): autoclave and put in dumpster Liquid waste: add disinfectant (bleach to 10%) and pour down drain
- 23. Autoclaves Autoclaves use pressurized steam to sterilize materials. There is usually steam remaining in the chamber at the end of a liquid cycle.
- 24. Autoclave Safety Opening door at end of liquid cycle: Wear eye and face protection. Stand behind door when opening it. Slowly open door only a crack to allow residual steam to escape. Keep face away from door as it opens.
- 25. Autoclave Safety Removing liquids at end of cycle: Wait 5 min. before removing liquids. Liquids removed too soon may be super-heated and boil up and out of container. Aim mouth of flask away from face. Don’t knock flask against bench.
- 26. Standard Microbiological Practices Decontaminate work surfaces daily and after any spill of viable material Report accidents to the PI Tell Health Care Provider that you work with infectious agents or chemicals
- 27. THANK YOU
Editor's Notes
- Biosafety is the section of Environmental Safety that addresses working safely in the laboratory with microorganisms and rDNA (organisms whose genetic material has been altered). Its goal is to help prevent lab-acquired infections while allowing researchers to work with organisms of various levels of hazard. This is done by advising researchers of the containment practices that are appropriate for the organisms they are handling, based on the level of risk presented by the organism.
- Various regulations and guidelines have been developed to protect workers and the environment. They define work practices required to work safely with biological material of all levels of hazard. All Principal Investigators must register their experiments that involve the use of rDNA, infectious microorganisms, and human blood, unfixed human tissue, and primary human cell lines with DES.
- There are 4 levels of biological containment that allow laboratory researchers to work safely with infectious agents of various levels of risk. They are designed to prevent lab-associated infections. BL1 is the lowest level of containment. At UM there are many BL1 labs, most of which are molecular biology labs that use non-pathogenic strains of E. coli for cloning. .
- BL2 is used for infectious agents that are transmitted by contact, ingestion, or puncture with contaminated sharp object. There are several BL2 labs at UM.
- BL2 labs are similar to BL1 laboratories, with the addition of these specific practices.
- BL3 containment is used for work with infectious agents that are transmitted by inhalation. We have 2 BL3 labs at UM, but neither is currently in use.
- There are only a very few BL4 labs in the country: CDC in Atlanta; NIH and Ft. Detrick in MD. BL4 is used for work with the most hazardous microorganisms. Hot Zone described BL4 containment.
- The OSHA BBP standard was published in 1991 to address concerns over occupational transmission of HIV to healthcare workers. BBPs are microorganisms present in the blood of infected persons, which are transmitted by blood-to-blood contact. Not casual contact or by inhalation. If you work with human material, you are covered by the BBP Standard.
- If you work with human material, OSHA requires: annual training of employees. If you work with human material, please take the interactive web based training available on our web page. provision of HBV vaccine by employer at no cost to employee; use of Universal Precautions when handling human material treating ALL material as if it may contain BBPs
- Even if material has been tested for BBPs such as HIV and HBV, it is possible that the test was performed before a detectable level of antibodies were produced, and the virus may be present. Therefore, ALL human material should be treated as infectious. Even if it is from your lab partner, because you can’t tell if someone is infected. If you work with human material, please take the interactive web based training available on our web page. (Copy of page in handout).
- Routes of occupational transmission of BBP. Optional: Routes of transmission in the general population: Sexual, both homosexual and heterosexual Sharing needles among injecting drug users. Blood transfusions prior to screening of blood supply (1980s??). Mother to unborn child.
- These are the basic practices when working with any microorganisms, and are the foundation for BL1 containment. They are designed to prevent transmission by contact, ingestion, and puncture. These practices are appropriate for ALL laboratories that work with radioisotopes and chemicals as well as microorganisms.
- Gloves are not a substitute for hand washing.
- In your handouts there is an information sheet on handling sharps safely. Sharps containers are available from Chemistry Stores and scientific distributors such as Fisher and VWR. Sharps containers are usually red, puncture-resistant plastic. Do not overfill sharps containers. Please call DES for pick-up of filled sharps containers. They do not need to be autoclaved before pick-up.
- Most needle sticks occur when recapping needles. If you must recap the needle, use a one-handed technique. If you get stuck by a needle, wash the area with soap and water, then go to the University Health Center for follow-up as soon as possible. Anti-retroviral drugs are more effective if given within 2-3 hours after needle stick.
- Use only autoclavable bags in autoclave. Put bags of waste in stainless steel or polypropylene tray during autoclaving to contain any leaks. There is information on autoclave safety and Biological Waste Disposal in the handout.
- Inform your healthcare provided that you work in a laboratory, and describe the types of hazardous materials you handle.