Standard Operating Procedure (SOP)

Here’s a detailed Standard Operating Procedure (SOP) for the Calibration of a Coulter Counter:

1. Purpose

To provide a standardized procedure for the calibration of the Coulter Counter, ensuring accurate and reproducible cell count and size distribution measurements.

2. Scope

This SOP applies to all Coulter Counter units used in [Insert Department/Lab Name] for particle or cell counting.

3. Responsibility

• Laboratory Technician/Analyst: Responsible for performing and documenting the calibration procedure.

• Lab Supervisor/QC Head: Responsible for reviewing calibration data and approving the instrument for use.

• QA Officer: Ensures periodic calibration and maintains audit records.

4. Definitions

• Coulter Counter: An electronic instrument that counts and sizes particles suspended in an electrolyte by detecting changes in electrical resistance.

• Calibration: Adjustment or verification of an instrument to ensure measurements match reference standards within accepted tolerance levels.

5. Materials and Equipment

• Coulter Counter

• Certified Latex or Polystyrene Calibration Beads (traceable to NIST or equivalent)

• Diluent/Electrolyte Solution (e.g., Isoton)

• Clean sample vials/tubes

• Calibration log sheet

• Timer or stopwatch

• Micropipettes with sterile tips

• Gloves and lab coat

6. Frequency of Calibration

• Daily/Before Each Use: Verification with calibration beads

• Monthly: Full calibration and size verification

• Annually: External certification (if applicable)

7. Procedure

7.1 Pre-Calibration Checks

1. Turn on the Coulter Counter and allow the system to stabilize as per the manufacturer’s instructions.

2. Ensure the aperture tube is clean and properly mounted.

3. Confirm the electrolyte solution is freshly prepared and degassed if necessary.

4. Prime the system (flush with diluent) to remove air bubbles and contaminants.

7.2 Calibration Using Standard Beads

1. Prepare Calibration Sample

   • Vortex the calibration bead suspension to ensure uniformity.

   • Pipette the recommended volume (usually 10–100 µL) into a clean vial with diluent (as per bead manufacturer’s instructions).

   • Mix gently to avoid foam or bubbles.

2. Run Calibration Sample

   • Place the sample vial under the aperture tube.

   • Initiate the count using the system software or interface.

   • Observe the peaks in the histogram for particle size and count.

3. Evaluate the Calibration Curve

   • Confirm that the peak size matches the certified mean diameter of the beads (within ±2%).

   • Confirm the count reproducibility by running the standard at least three times and calculating the %CV (should be ≤5%).

4. Adjust Settings (if applicable)

   • Adjust gain, threshold, or calibration factors based on software prompts to match bead specifications.

   • Save settings as “calibrated” when performance is within acceptance criteria.

7.3 Recording and Verification

• Record:

  • Bead type and lot number

  • Measured size and count values

  • Instrument settings

  • Deviation (if any) and corrective action taken

• Print or save calibration histogram/results for documentation.

8. Acceptance Criteria

9. Post-Calibration Steps

• Rinse the system with diluent after calibration.

• Label the Coulter Counter as “Calibrated” with date and next due date.

• If the calibration fails:

  • Do not use the instrument for analysis.

  • Report the issue to maintenance or service provider.

  • Record all deviations and actions taken.

10. Documentation and Records

Maintain the following:

• Daily and monthly calibration logs

• Calibration certificate of standard beads

• Histogram printouts or digital records

• Service and maintenance records (if calibration fails)

11. Safety Precautions

• Wear appropriate PPE (gloves, lab coat, eye protection).

• Use only approved calibration beads and diluents.

• Do not operate the instrument with cracked apertures or leaks.

12. References

• Instrument Manufacturer’s User Manual

• ISO 13319: Determination of particle size distribution — Electrical sensing zone method

• In-house Quality Control Manual.

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