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SOP-012: Operation of a Class II Biosafety Cabinet

Purpose​

To ensure the safe and aseptic operation of a Class II Biosafety Cabinet (BSC) during mammalian cell culture procedures.

Scope​

This SOP applies to all personnel performing cell culture work in a certified Class II Biosafety Cabinet.

Responsibilities​

The operator is responsible for:

  • Verifying the cabinet is functioning properly before use.
  • Maintaining aseptic workflow.
  • Preventing disruption of cabinet airflow.
  • Cleaning and disinfecting the cabinet before and after use.

Required Materials​

Reagents​

  • 70% ethanol
  • Appropriate disinfectant (if required by institutional policy)

Equipment​

  • Certified Class II Biosafety Cabinet
  • Sterile laboratory consumables
  • Biohazard waste container
  • Laboratory wipes

Procedure​

1. Pre-Operation Checks​

Confirm that:

  • The biosafety cabinet certification is current.
  • The airflow alarm is not activated.
  • The sash is positioned at the recommended operating height.
  • The work surface is clean and free of unnecessary materials.

2. Prepare the Cabinet​

Turn on the cabinet according to laboratory procedures.

Allow the cabinet to operate for the recommended stabilization period before beginning work.

Disinfect the work surface with 70% ethanol.

Arrange materials to maintain an organized workflow.

3. Perform Cell Culture Procedures​

Work well within the interior work area.

Keep materials organized from clean to contaminated.

Avoid blocking the front or rear air grilles.

Perform slow, deliberate movements to minimize airflow disruption.

Minimize unnecessary entry and exit of hands and materials.

4. Complete the Procedure​

Seal all culture vessels.

Remove waste and contaminated materials.

Disinfect all reusable equipment before removal.

Disinfect the work surface thoroughly.

Allow the cabinet to continue operating according to laboratory policy before shutting it down, if applicable.

Acceptance Criteria​

ParameterAcceptance Criterion
AirflowUnobstructed throughout operation
Work surfaceClean before and after use
Cabinet organizationMinimal clutter
Cabinet statusNo alarms or operational faults

Critical Control Points​

  • Maintain the sash at the recommended operating height.
  • Do not block front or rear air grilles.
  • Avoid rapid arm movements inside the cabinet.
  • Keep only essential materials inside the work area.
  • Separate clean and contaminated materials throughout the procedure.

Documentation​

Record the following when required:

  • Date
  • Operator
  • Procedure performed
  • Cabinet identification number
  • Any equipment malfunction or abnormal observations

Quality Control​

Verify:

  • Cabinet certification is current.
  • Airflow indicators function normally.
  • Cabinet surfaces are properly disinfected.
  • Cabinet is free of unnecessary materials.
  • Waste has been removed following the procedure.

Troubleshooting​

ObservationPossible CauseCorrective Action
Airflow alarm activatedObstructed airflow or cabinet malfunctionStop work and investigate before continuing
Excessive clutterPoor workspace organizationRemove unnecessary materials from the cabinet
Frequent contaminationImproper aseptic workflowReview cabinet practices and airflow management
Difficulty maintaining sterilityExcessive movement inside cabinetWork slowly and maintain proper workflow

Safety Considerations​

  • Wear appropriate personal protective equipment (PPE).
  • Never use the cabinet if airflow or safety alarms indicate malfunction.
  • Do not use open flames inside the biosafety cabinet.
  • Dispose of biological waste according to institutional biosafety procedures.
  • Report cabinet malfunctions immediately.