Working with Biological Working with Biological Materials in the Materials in the

O’Connell LaboratoryO’Connell Laboratory

Grace OGrace O’’Connell, PhD Connell, PhD

[Date Training Is Given][Date Training Is Given]

Basic RequirementsBasic RequirementsResearch involving biological agents is Research involving biological agents is regulated by the National Institutes of regulated by the National Institutes of Health (NIH), Centers for Disease Control Health (NIH), Centers for Disease Control and Prevention, the US Department of and Prevention, the US Department of Agriculture and the State of California. Agriculture and the State of California. UC Berkeley requires a BUA approved by UC Berkeley requires a BUA approved by CLEB prior to the initiation of the CLEB prior to the initiation of the experiment. CLEB works with campus experiment. CLEB works with campus Biosafety Officer Bob Hashimoto, who Biosafety Officer Bob Hashimoto, who can provide additional guidance (can provide additional guidance (bobhash@berkeley.edu or 643-6562).

Purpose of This TrainingPurpose of This Training

UC Berkeley and CLEB require that the UC Berkeley and CLEB require that the PI provide lab-specific biosafety PI provide lab-specific biosafety training on the hazards of the specific training on the hazards of the specific experimental procedures used in their experimental procedures used in their laboratory to members of their laboratory to members of their research group. One way to document research group. One way to document this training is to record it in the this training is to record it in the minutes of a group meeting, listing all minutes of a group meeting, listing all participants.participants.

Contents of This TrainingContents of This Training

• CLEB Administrative RequirementsCLEB Administrative Requirements• Role of the Investigator/Lab StaffRole of the Investigator/Lab Staff• Description of the Experiment-BUADescription of the Experiment-BUA

– The Agents UsedThe Agents Used– The TemplateThe Template

• Risk Assessment/Biosafety LevelRisk Assessment/Biosafety Level– Good Work PracticesGood Work Practices– Personal Protective EquipmentPersonal Protective Equipment– Engineering ControlsEngineering Controls

• Waste Disposal/DecontaminationWaste Disposal/Decontamination• Emergency ProceduresEmergency Procedures

CLEB Administrative CLEB Administrative RequirementsRequirements

• CLEB requires the following from every CLEB requires the following from every PI approved to use biological agents:PI approved to use biological agents:– Train all research group members on lab-Train all research group members on lab-

specific hazards and proceduresspecific hazards and procedures– Report incidents involving biological agents Report incidents involving biological agents

and clinical specimens immediately to EH&Sand clinical specimens immediately to EH&S– Submit a BUA amendment for any change in Submit a BUA amendment for any change in

personnel or Scope of Work personnel or Scope of Work

Description of the Experiment-Description of the Experiment-BUABUAThe Scope of Work-from the OThe Scope of Work-from the O’’Connell BUA Connell BUA narrativenarrative• The scope of the research includes mechanical The scope of the research includes mechanical

testing of animal and human samples and cell testing of animal and human samples and cell culture. culture.

• Spine sections will be acquired either from a local Spine sections will be acquired either from a local abattoir or through a donor bank (e.g. NDRI for abattoir or through a donor bank (e.g. NDRI for human samples). human samples).

• Care should be taken when using equipment for Care should be taken when using equipment for sample preparation and testing. Equipment sample preparation and testing. Equipment should be considered as biohazard contaminated should be considered as biohazard contaminated (performed in 2166 Etcheverry).(performed in 2166 Etcheverry).

Description of the Experiment-Description of the Experiment-BUABUAThe Agents UsedThe Agents Used

The OThe O’’Connell lab uses the following:Connell lab uses the following:

•Human specimensHuman specimens

•Animal specimens (bovine)Animal specimens (bovine)

•Human CellsHuman Cells

Description of the Experiment-Description of the Experiment-BUABUAThe Risks That May Affect Containment The Risks That May Affect Containment Level Level These are some of the common experimental These are some of the common experimental risks associated with our research: risks associated with our research: •The primary route of exposure of our biological materials is The primary route of exposure of our biological materials is through the bloodborne route. Procedures involving sharps, through the bloodborne route. Procedures involving sharps, especially when handling human and animal subjects is of especially when handling human and animal subjects is of primary concern. primary concern.

•All employees and students are encouraged to receive All employees and students are encouraged to receive vaccination against hepatitis B.vaccination against hepatitis B.

•All used sharps should be placed in rigid hard plastic sharps All used sharps should be placed in rigid hard plastic sharps containers that will be brought to the red bag consolidation area containers that will be brought to the red bag consolidation area for disposal as medical waste.for disposal as medical waste.

•Reusable metal sharps shall be autoclaved at 121 degrees C for Reusable metal sharps shall be autoclaved at 121 degrees C for 30 minutes and 20 PSI.30 minutes and 20 PSI.

Description of the Experiment-Description of the Experiment-BUABUAThe Risks That May Affect Containment The Risks That May Affect Containment Level Level • All human clinical specimens shall be handled with standard

Universal Precautions. • Universal PrecautionsUniversal Precautions is an approach to infection control. is an approach to infection control.

According to the concept of Universal Precautions, all human According to the concept of Universal Precautions, all human blood and certain human body fluids are treated as if known blood and certain human body fluids are treated as if known to be infectious for HIV, HBV, and other bloodborne to be infectious for HIV, HBV, and other bloodborne pathogens. pathogens.

• Our lab will perform analytical and tissue culture studies with human and animal specimens.

• You should apply 10% bleach and 70% ethanol as disinfectants to clean up following an experiment.

Risk Assessment/Biosafety Risk Assessment/Biosafety LevelLevelSummary of Good Work PracticesSummary of Good Work Practices

• Wash Hands FrequentlyWash Hands Frequently

• Use Personal Protective Equipment Use Personal Protective Equipment and Engineering Controlsand Engineering Controls

• Minimize Use of Sharps and if used, Minimize Use of Sharps and if used, handle carefullyhandle carefully

• Do Not Eat, Drink, or Smoke in the labDo Not Eat, Drink, or Smoke in the lab

• Disinfect Work AreasDisinfect Work Areas

Risk Assessment/Biosafety Risk Assessment/Biosafety LevelLevelGood Work Practices Good Work Practices

• Centrifugation:Centrifugation: During cell During cell preparation, samples will need to be preparation, samples will need to be centrifuged in a sealed vial (the centrifuged in a sealed vial (the centrifuge will contain a cone and centrifuge will contain a cone and will be self contained). will be self contained). – Transport samples in sealed tubes Transport samples in sealed tubes

placed in a tube rack. Care will be taken placed in a tube rack. Care will be taken to avoid spillage.to avoid spillage.

Risk Assessment/Biosafety Risk Assessment/Biosafety LevelLevelGood Work Practices Good Work Practices

• Sample preparation:Sample preparation: Spine Spine samples will need to be dissected for samples will need to be dissected for biomechanical studies. biomechanical studies. – Proper safety attire shall be worn, Proper safety attire shall be worn,

including gloves, lab coat, safety including gloves, lab coat, safety goggles, and appropriate clothinggoggles, and appropriate clothing

• Never work in the laboratory alone. Never work in the laboratory alone.

Risk Assessment/Biosafety Risk Assessment/Biosafety LevelLevelGood Work Practices Good Work Practices

• Room 5102 is additional wet lab space to perform Room 5102 is additional wet lab space to perform assays. assays.

• Sample TransportationSample Transportation: When transporting : When transporting samples between rooms, make sure solutions are samples between rooms, make sure solutions are sealed and placed in a hard sided container.sealed and placed in a hard sided container.

• Samples should be fixed with 4% paraformaldehyde Samples should be fixed with 4% paraformaldehyde or previously frozen at -20or previously frozen at -20ooC prior to transportation. C prior to transportation. Fixing and freezing are effective methods make the Fixing and freezing are effective methods make the tissues no longer viable. tissues no longer viable.

• All solutions will be labeled and biohazard solutions All solutions will be labeled and biohazard solutions will be labeled with a biohazard sticker.will be labeled with a biohazard sticker.

Risk Assessment/Biosafety Risk Assessment/Biosafety LevelLevelGood Work Practices Good Work Practices

• Autoclave: Autoclave: Tools that will need to be used Tools that will need to be used sterilely will be autoclaved. These items include sterilely will be autoclaved. These items include medical tools, glass bottles, and pipette tips. medical tools, glass bottles, and pipette tips.

• Note: Note: Biohazard waste should be disposed of in Biohazard waste should be disposed of in red biohazard bags and disposed of through red biohazard bags and disposed of through EH&S. Biohazard waste should not be autoclaved.EH&S. Biohazard waste should not be autoclaved.

• Caution: Caution: Autoclaves can be a physical hazard Autoclaves can be a physical hazard due to the heat and steam so no one is permitted due to the heat and steam so no one is permitted to use it unless they have been trained on the to use it unless they have been trained on the safety procedures and how to operate the safety procedures and how to operate the settings.settings.

Risk Assessment/Biosafety Risk Assessment/Biosafety Level Level Personal Protective EquipmentPersonal Protective Equipment

• The following personal protective The following personal protective equipment must be worn when equipment must be worn when manipulating tissue or using the manipulating tissue or using the biosafety cabinet: biosafety cabinet: – Gloves (latex or nitrile)Gloves (latex or nitrile)– Water resistant lab coatWater resistant lab coat– Goggles/safety glasses Goggles/safety glasses

Risk Assessment/Biosafety Risk Assessment/Biosafety Level Level Engineering ControlsEngineering Controls

• The following engineering controls must be The following engineering controls must be used whenever handling tissue in the used whenever handling tissue in the laboratory:laboratory:– Biosafety cabinetBiosafety cabinet– Sink for Hand WashingSink for Hand Washing– Mechanical pipettesMechanical pipettes– Sealed vialsSealed vials– Sharps containers/Broken Glass boxSharps containers/Broken Glass box– Eyewash/deluge shower (use only in an Eyewash/deluge shower (use only in an

emergency)emergency)

Waste Waste Disposal/DecontaminationDisposal/Decontamination

• Waste Disposal: Waste Disposal: Tissue culture plastic used for cell Tissue culture plastic used for cell culture will be bleached then disposed of in a biohazard culture will be bleached then disposed of in a biohazard bag. Red bag waste shall be picked up by Environmental bag. Red bag waste shall be picked up by Environmental Health and Safety. Health and Safety.

• Paraformaldahyde and any mixed biological and chemical Paraformaldahyde and any mixed biological and chemical waste solutions should be disposed of through EH&S as waste solutions should be disposed of through EH&S as chemical waste.chemical waste.

• Spill procedures Spill procedures (biological tissues/blood/waste)(biological tissues/blood/waste): : Clean Clean spills by soaking it in 10% bleach for 20 minutes before spills by soaking it in 10% bleach for 20 minutes before cleaning up and disposing as biohazardous waste.cleaning up and disposing as biohazardous waste.– Notify Dr. O’Connell and EH&S of any spills within 8 hours.Notify Dr. O’Connell and EH&S of any spills within 8 hours.

Emergency ProceduresEmergency Procedures

• In the event of a laboratory accident:In the event of a laboratory accident:– Tend to injured person, wash the wound Tend to injured person, wash the wound

with water.with water.– Notify Dr. O’Connell or the lab supervisor Notify Dr. O’Connell or the lab supervisor

IMMEDIATELY so that CLEB and the IMMEDIATELY so that CLEB and the Biosafety Officer can be notified within 8 Biosafety Officer can be notified within 8 working hours. working hours.

– Initiate clean up procedures.Initiate clean up procedures.– Notify EHS, if the PI is not available, 642-Notify EHS, if the PI is not available, 642-

3073 or 642-3333 (After Hours)3073 or 642-3333 (After Hours)– Seek medical attention at Tang or the Alta Seek medical attention at Tang or the Alta

Bates ER at 2450 Ashby Avenue after Bates ER at 2450 Ashby Avenue after hours. hours.

ConclusionConclusionThis training is designed to inform you of the This training is designed to inform you of the regulatory requirements regarding use of regulatory requirements regarding use of biological materials and our labbiological materials and our lab’’s s implementation of the campus safety implementation of the campus safety requirements. requirements.

All experiments with human clinical specimens All experiments with human clinical specimens by CLEB and any changes require an by CLEB and any changes require an approved amendment to the BUA.approved amendment to the BUA.

Contact the Biosafety Officer for assistance, if Contact the Biosafety Officer for assistance, if needed (Bob Hashimoto). needed (Bob Hashimoto).