Biological Safety Training

Introduction to Biological Safety Biological Hazards are divided into 4 Biosafety Levels BSL 1 BSL 2 BSL 3 BSL4 Biosafety levels define the lab requirements, protective clothing, and work practices

BioSafety Definitions BSL 1 (BMBL) Not known to consequently cause disease in healthy human adults Pose minimal hazards under ordinary conditions of handling

BioSafety Level 1 Standard Practices Use Mechanical Pipetting devices No Eating, Drinking, Smoking in Lab Minimize splashes and aerosols Decontaminate work surfaces Safe handling of sharps Wash Hands before leaving lab

BioSafety Level 1 Protective Clothing Lab Coat Gloves Eye Protection

BioSafety Definitions BSL 2 (BMBL) suitable for work involving well-characterized agents not known to cause disease in healthy adult humans, and of minimal potential hazard to laboratory personnel and the environment Examples: Measles Virus, Salmonella, Hepatitis B Virus

BioSafety Definitions Other Materials Handled at BSL 2 (Requires compliance with OSHA BBP Standard) Human Blood Human Blood Components Human Tissues Human Cell Lines

BioSafety Level 2 Standard Practices All requirements for BSL 1 plus: Access to laboratory is limited or restricted when work is being conducted Personnel have specific training in handling pathogenic agents Biohazard Sign posted on the door Extreme precautions are taken with contaminated sharp items

BioSafety Level 2 Standard Practices cont. Written Biosafety Procedures Report Spills Minimize aerosol generation Personnel receive appropriate immunizations or test (e.g. Hep B vaccine or TB skin Testing)

BioSafety Level 2 Written Biosafety Procedures Prepared specifically for lab by PI Incorporated into Lab Safety Plan Personnel are advised of special hazards Personnel are required to read and follow procedures

BioSafety Level 2 Personal Protective Equipment Lab Coats Gloves Double Gloving when appropriate Eye Protection - Safety Glasses/Shield

BioSafety Level 3 BSL 3 (BMBL) applicable to clinical, diagnostic, teaching, research, or production facilities in which work is done with indigenous or exotic agents which may cause serious or potentially lethal disease as a result of exposure by the inhalation route.

BioSafety Level 3 Standard Practices All requirements for BSL 2 plus: Limited lab access 2 doors in a series to access lab Able to wash entire lab Special exhaust ventilation (Not re circulated, negative pressure)

BioSafety Level 3 Standard Practices –cont. High level of training Personnel receive vaccinations if available Work in Biosafety cabinets

BioSafety Level 3 Examples Mycobacterium Tuberculosis Hanta virus SARS

BioSafety Level 4 BSL 4 (BMBL) required for work with dangerous and exotic agents which pose a high individual risk of aerosol-transmitted laboratory infections and life-threatening disease.

BioSafety Level 4 Requirements All requirements for BSL3 plus: Class III Biosafety cabinet or positive pressure suits Shower/Change rooms Clothing Autoclaved before laundering Air Locks

BioSafety Level 4 Examples Ebola Virus Monkey B Virus Marburg Virus

Containment Primary Protection of personnel and immediate lab from exposure Good Techniques Safety equipment Biological Safety Cabinet Engineering Controls PPE Serologic surveillance

Containment Secondary Protection of environment external to lab Facility Design Operational practices

Safe Work Practices for all Levels Wash hands after work; when removing gloves; before leaving lab No eating, drinking, applying cosmetics, handling contact lenses in the lab. Maintain labs in clean, orderly fashion.

Safe Work Practices for all Levels Limit access to lab when work with organisms is in progress Use good microbiological techniques No mouth pipetting When possible use plastic instead of glass

Sharps Safety Sharps include needles, syringes, razor blades, lancets, slides, scalpels, pipettes, micropipettes, pipette tips, broken plastic or glassware, and other devices capable of cutting or piercing the skin.

Sharps Safety Contaminated needles shall not be bent, recapped, or removed unless there is no feasible alternative. If required, use a mechanical device or a one handed technique.

Sharps Safety Safety devices or alternatives to needles should be used when available.

Sharps Disposal Sharps containers for disposal of these items should be conveniently located and easily accessible in all work places where sharps are used.

Sharps Disposal Syringes and syringes without a needle attached go into a sharps container Contaminated micropipettes, pipette tips, and Pasteur pipettes are discarded in a puncture-resistant container or a sharps container for disposal.

Sharps Disposal Don’t place needles or sharps in office waste containers

Laminar Flow Equipment BioSafety Cabinets (BSCs) contain infectious agents to protect personnel and the environment Laminar Flow Clean Benches (LFBs) Non Hazardous work only Protect work from contamination

BioSafety Cabinets 3 Classes All exhaust is HEPA filtered before leaving the cabinet Class I Do not protect the work from contamination Air entering cabinet is not filtered

Biosafety Cabinets Class II (4 types – A, B1, B2, and B3) Each type recirculates different amount of air Some are hard ducted, and some exhaust into the room. Class III Totally enclosed, ventilated cabinets Work through portals with attached gloves

BioSafety Cabinets All equipment is laid out to not restrict airflow in the cabinet

Laminar Flow Benches Do not protect personnel or the environment Discharges HEPA filtered air across the work surface toward the user.

Use of Laminar Flow Equipment Minimize airflow disturbances (moving in/out of cabinet, people walking by, opening doors, blocking grills with equipment) Wipe down surfaces with approved disinfectant before and after work.

Use of Laminar Flow Equipment Minimize use of flames in cabinet Try not to use chemicals in Class I or II hoods that are ducted Do not store excess equipment in cabinet All laminar flow equipment must be certified annually.

Centrifuges

Centrifuge Hazards Mechanical failure of Machine Lab equipment failure (tubes etc.) Aerosol generation Operator error

Centrifuge Operation Check tubes for cracks/chips Use matched sets of tubes, buckets, etc. Make sure the rotor is balanced properly Tightly seal all tubes and safety cups Ensure that rotor is locked to spindle and the bucket is seated Close lid during operation Allow to come to complete stop before opening

Centrifuge Safety Use sealed tubes, safety buckets or rotors when possible When possible fill and open centrifuge tubes or buckets in the biological safety cabinet. Lubricate O-rings and rotor threads weekly Disinfect weekly and after all spills or breaks

Biohazardous Waste All cultures, stocks, and other regulated waste are decontaminated before disposal by an approved decontamination method such as autoclaving.

Biohazardous Waste Examples stock cultures materials containing or contaminated with blood blood and body fluids sharps pipette tips animal waste other contaminated lab material

Blood and Body Fluids Waste Serum, Plasma Other blood components Semen Vaginal secretions Cerebrospinal fluid Pleural fluid Peritoneal fluid Pericardial fluid Amniotic fluid Any other body fluid visibly contaminated with blood

Infectious Sharps Waste Hypodermic needles Scalpels Pipettes /pipette tips Breakable containers Glass products (i.e., slides or cover skips)

Biohazardous Waste Inappropriate materials Dispose of noninfectious waste in regular trash

Biohazardous Waste Inappropriate materials These include: aerosol cans, chemical waste, flammable material

Biohazardous Waste Inappropriate packaging Use puncture resistant package if needed

Biohazardous Waste Containers Biohazardous waste containers shall be clearly marked with the universal biohazard symbol.