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Biosafety

Biosafety levels and Cabinet in laboratory

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Biosafety

  1. 1. Dr. Malathi M 20/11/2014
  2. 2. Topics to be discussed : • What is biosafety? • Why we need ? • Levels of biosafety • Biosafety cabinet • Types • Decontamination
  3. 3. Biosafety is the application of safety precautions that reduce a laboratorians risk of exposure to a potentially infectious material and limit contamination of the work environment and ultimately the community { CDC }
  4. 4. Why we need biosafety ???? 1. Lab has hazards of processing infectious agents 2. Accidental threat to workers and environment 3. To have adherence with safety regulations while dealing with highly infectious agents
  5. 5. BIOSAFETY LEVEL 1 • Microbes not known consistently to cause disease in healthy adults and present minimal potential hazard to lab and environment • Eg : non pathogenic strain of E.coli
  6. 6. BSL – 1 practices: • Standard microbiological practices are followed • Work can be performed on an open table or bench • PPE{Personal protective equipment} needed • Sink – hand washing • Lab – doors seperate
  7. 7. Biosafety level 2 • Microbes that possess moderate hazards to laboratorians • Eg: Staphylococcus aureus
  8. 8. BSL – 2 practices: • Access to lab is restricted when work is being conducted • PPE , face shields, eye goggles • Biosafety cabinet • Autoclave/Decontamination proper • Self closing doors • Sink with eyewash apparatus readily available
  9. 9. Biosafety level 3 • Serious / potentially lethal disease through respiratory transmission • Eg: Mycobacterium tuberculosis
  10. 10. BSL 3 - practices • Laboratorians – under medical surveillance and receive immunisation • Access to lab restricted & controlled • PPE with respirators • BSC • Sink with eyewash • Exhaust air – not recirculated • Self closing doors with automatic locking
  11. 11. Biosafety level 4 • Highest level of biological safety • Dangerous and exotic microbes • Eg: Ebola , marburg viruses
  12. 12. BSL 4
  13. 13. BSL 4 - practices • Change clothes before entering • Shower upon exiting • Decontaminate all materials before exiting • Class III BSC • Separate building for lab • Vacuum lines and decontamination systems
  14. 14. BSL 4 lab
  15. 15. Introduction • Biosafety cabinets (BSCs) are primary means of containment, developed for working safely with infectious micro-organisms • BSCs are only one overall part of biosafety program, which requires consistent use of – good microbiological practices – primary containment equipment – primary containment facility design
  16. 16. To be precise, “BSCs are designed to provide personnel, environmental and product protection when appropriate practices and procedures are followed” Adapted from CDC-BMBL- 5th Edition/1999 Appendix A – Primary Containment for Biohazards: Selection, Installation and Use of Biological Safety Cabinets
  17. 17. Historical perspective 1. Early prototype clean air cubicles (clean filtered air was blown directly at the working surface inside a cubicle – this places the personnel in a contaminated air stream) 2. Concept of small workstation (non-ventilated cabinets – wood/stainless steel) 3. Ventilated cabinets (lack of controlled/ adequate air flow leading on to mass airflow) Class I 4. HEPA filter were introduced (undergoing modifications till date)
  18. 18. • HEPA – High efficiency particulate air filter • It removes the most penetrating particle size (MPPS) of 0.3 μm with an efficiency of at least 99.97 % • The typical HEPA filter is a single sheet of borosilicate fibers treated with a wet-strength water-repellant binder
  19. 19. • The filter medium is pleated to increase the overall surface area, with pleats being separated by corrugated aluminum tubes • This separation is mainly to prevent collapse • It removes particulate matter by three mechanisms interception, impaction, diffusion • The filtering efficiency depends upon fiber diameter, filter thickness and face velocity • These filters are fitted either in the exhaust or air supply system to remove particulate matter
  20. 20. http://en.wikipedia.org/wiki/HEPA
  21. 21. Importance of a Biosafety cabinet • Provide protection to the – personnel handling infectious material – environment by preventing the release of microbes – product (e.g. in handling cell cultures)
  22. 22. BIOSAFETY CABINET - I • Provides personnel and environmental protection, but no product protection • Exhaust system – HEPA filter • Class I BSC – unfiltered room air is drawn in through the work opening and across the work surface • Inward airflow – Minimum velocity – 75 linear feet / minutes
  23. 23. • To enclose equipment (Eg. Centrifuges, harvesting equipment, small fermenters) • For procedures with potential to generate aerosols ( tissue homogenation, culture aeration) • Class I BSC is hard-ducted • Cabinet air is drawn through a HEPA filter as it enters the cabinet exhaust plenum.
  24. 24. REQUIREMENTS: • Open fronted • Glass in the upper front • An integral tray to contain spills and splashes • Inward airflow – 0.7 to 1 m/sec • Protection factor – 1.5 * 105 • Protection factor = number of particles which, if liberated into the air of the cabinet will not escape into the room • Filtration from the exhaust air - HEPA
  25. 25. CLASS I cabinet
  26. 26. Biosafety cabinet class II • Product protection • Predictable particle behaviour • Laminar air flow principle (1960) • Particle barrier systems • Risk of contaminant release into the lab and risk of product contamination
  27. 27. Class II A1 A2 B1 B2
  28. 28. CLASS II – Type A1 • Internal fan – draws room air – 75lfm velocity • Supply air flows through HEPA – particulate free air to the work surface • Reduced turbulence • Reduced cross contamination
  29. 29. • Downward moving air – splits into two 1) To the front grille 2) To the rear grille • 30% of the air – exhaust HEPA filter • 70% of the air – recirculates through HEPA filter back into the work zone of the cabinet
  30. 30. • Not to be used for work involving volatile toxic chemicals • Exhaust the air outside the building ( through use of canopy hood and filter housing ) • CLASS II A1 and A2 – never be hard ducted to the building exhaust system
  31. 31. CLASS II A1
  32. 32. CLASS II A2 ( formerly B3) • Inflow air velocity 100lfm • all positive pressure contaminated plenums within the cabinet are surrounded by a negative air pressure plenum  ensures leakage
  33. 33. CLASS II A2
  34. 34. CLASS II B1 • For hazardous chemicals and carcinogens • Designed and originated with the National cancer institute type 212 ( later called Type B) • Definition of Type B1 cabinets: • Classic NCI design Type B, and cabinets without supply HEPA filters located immediately below the work surface, or those with exhaust/recirculation down flow splits other than exactly 70/30%
  35. 35. • Cabinet supply blowers draw room air through the front grille and through HEPA • Inflow velocity 100lfm • Split in the down flowing air stream just above the work surface • 70% air  through the rear grille  exhaust HEPA filter  discharge through building • 30% air  Down flow air  front grille
  36. 36. CLASS II B1
  37. 37. CLASS II B2 • Total exhaust cabinet • No air recirculation • Simultaneous biological and chemical containment • Inflow air velocity 100lfm • Exhaust 1200 cubic feet/min of room air  expensive cabinet high cost of heavier gauge and higher capacity exhaust fan  hence only for research
  38. 38. CLASS II B2
  39. 39. CLASS III • Highly infectious agents, hazardous operations • Gas tight  no leak greater than 1 *10-7 cc/sec with 1% test gas at 3 inches pressure water gauge • Non opening view window • Passage of materials through a dunk tank • Double door pass through box with autoclave
  40. 40. • Supply and exhaust air  HEPA • Negative pressure cabinet • No exhaust through the general lab exhaust • Long heavy duty rubber gloves attached in a gas tight manner to port in the cabinets
  41. 41. Class III
  42. 42. Work practices and procedures • Checklist of materials and work activity protocol • Arm movement slowly • Minimum persons • Lab coats buttoned fully • Proper Stool height
  43. 43. Check list • Daily check of airflow by airflow indicator and monthly or weekly with an anemometer • Ideal air flow – 0.7 to 1 m/s
  44. 44. • All procedures should be done atleast four inches in from the front grille • Only the materials needed for work should be kept inside • Wait for minimum of four minutes to switch off the blowers after the work is over
  45. 45. Decontamination • Disinfectant selection  EPA registration number in the label and list of infectious agents that the disinfectant is effective • BSC – ethanol not used as decontamination as it evaporates – no proper contact time – ethanol can be used as a rinsing agent • Formaldehyde vapour sterilisation to be done to kill spores
  46. 46. Disinfection method A • Cabinets with an internal electric power supply • Place 25 ml formalin(cabinet with internal volume of 0.38cu.m) to a vaporizer, or into a beaker on a hotplate • Close the cabinet and ensure that the exhause blow back valve is closed • Boil away formalin
  47. 47. Disinfection method B • 35ml formalin in a 100ml beaker inside the cabinet  add 10g potassium permanganate  seal the cabinet • Leave the cabinet at least 5 hours , preferably overnight and label DANGER – FUMIGATION IN PROGRESS • Open next day and work after 30 min for residual formaldehyde to exhaust
  48. 48. • Use of Ultraviolet lamps for BSC is not advisable { NIH, CDC }
  49. 49. Summary Class I Class III Class II
  50. 50. References • Appendix A – primary containment for biohazards – CDC article • Koneman`s color atlas and T.B of diagnostic microbiology • Diagnostic microbiology – Bailey and scott – 13th edition • Practical medical microbiology – Mackie and Mccartney – 14th edition
  51. 51. Thank you……….
  52. 52. Questions pls

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