SOP FOR ANALYST QUALIFICATION FOR MICROBIOLOGICAL ANALYSIS

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SOP FOR ANALYST QUALIFICATION FOR MICROBIOLOGICAL ANALYSIS

1.0 OBJECTIVE

To establish a standardized procedure for qualification and requalification of analysts performing microbiological analysis to ensure competency, accuracy, and compliance with regulatory requirements.

2.0 SCOPE

This SOP applies to all microbiologists working in the Microbiology Department involved in microbiological testing activities.

3.0 RESPONSIBILITY

3.1 Trainee Officer / Officer – Microbiology

  • Perform qualification activities as per this SOP.

  • Record observations and sign relevant documents.

  • Assist in preparation and updating of SOPs.

3.2 Senior Officer / Executive – Microbiology

  • Provide training to microbiologists.

  • Verify correct execution of procedures.

  • Review qualification documents and SOPs.

3.3 QC Head / Designee

  • Review overall qualification activities.

  • Approve qualification reports.

  • Ensure compliance with this SOP.

3.4 QA Head

  • Approve the SOP.

4.0 ACCOUNTABILITY

The Head of Department shall be accountable for effective implementation of this SOP.

5.0 DEFINITIONS & ABBREVIATIONS

5.1 Definition

Not Applicable (NA)

5.2 Abbreviations

SOP – Standard Operating Procedure
QC – Quality Control
QA – Quality Assurance
MID – Microbiology Department
IPA – Isopropyl Alcohol
BET – Bacterial Endotoxin Test
RSD – Relative Standard Deviation
LAF – Laminar Air Flow
LPC – Liquid Particle Counter
WFI – Water for Injection

6.0 PRECAUTIONS, SAFETY & EHS

6.1 Wipe sample bottles/vials with filtered 70% IPA before transferring into sterility area.
6.2 Autoclave all materials used in sterility operations.
6.3 Flame the neck of vials properly before opening.
6.4 Perform microbial testing under aseptic conditions to prevent contamination.
6.5 Incubate tubes in a stable heating block; avoid vibration. Do not incubate fragile gel tubes inside LAF cabinets.

7.0 MATERIALS & EQUIPMENT REQUIRED

7.1 For Bacterial Endotoxin Test (BET)

  • LAL Reagent

  • Control Standard Endotoxin (CSE) with COA

  • LAL Reagent Water (LRW)

  • Calibrated heating block

  • Calibrated micropipettes

  • Endotoxin-free tips

  • Depyrogenated glassware (10×75 mm, 12×75 mm, 16×100 mm)

  • Cyclo mixer

  • Test tube stand

7.2 For Particulate Matter Test

  • Particle-free WFI

  • Clean 100 ml vessel / beaker

  • 70% IPA

  • Liquid Particle Counter (LPC)

  • Computer system

7.3 For Sterility Testing

  • Sample to be tested

  • Vacuum pump & filtration assembly

  • Sterile FTM & SCDM media

  • Membrane filters (0.45 µm)

  • Diluting Fluid A (0.1% Peptone)

  • LAF unit

  • Sterile forceps and scissors

  • Bunsen burner

7.4 For Microbial Limit Test & Bioburden Test

  • LAF

  • Incubators (20–25°C & 30–35°C)

  • Culture media

8.0 PROCEDURE

8.1 Gowning Qualification

8.1.1 Production Analyst

Enter sterile production area following respective gowning SOP.

8.1.2 Microbiologist

  • Enter aseptic area as per Entry & Exit SOP.

  • Perform personal monitoring using contact plates from:

    • Forehead

    • Left & Right fingers

    • Left & Right booties

  • Incubate plates as per requirement.

  • Perform monitoring for three consecutive days.

  • Record results.

8.2 Bacterial Endotoxin Test (BET) Qualification

8.2.1 Select three recently approved batches of same product.
8.2.2 Analyze each batch individually as per BET SOP.
8.2.3 Compare results with historical data.

Invalid Test Conditions:

  • λ/4 concentration does not show positive result.

  • 2λ concentration does not show negative result.

  • Positive result in negative control.

Acceptance Criteria:

  • λ/4: Negative in all replicates

  • λ and 2λ: Positive in all replicates

  • λ/2: May or may not be positive

Record results in BET Qualification Format.
Upon successful qualification, issue Analyst Qualification Certificate.

8.3 Particulate Matter Qualification

8.3.1 Collect WFI in 100 ml clean vessel.
8.3.2 Perform testing using LPC as per SOP.
8.3.3 Conduct test for three consecutive days.

Acceptance Criteria:

  • ≤ 6000 particles ≥10 µm

  • ≤ 600 particles ≥25 µm

Record results in qualification format.
Issue qualification certificate after successful completion.

8.4 Sterility Test Qualification

8.4.1 Prepare 60 vials (100 ml WFI each).
8.4.2 Autoclave at 121°C for 30 minutes (validated cycle).
8.4.3 Divide into three groups (A, B, C).
8.4.4 Spike selected vials (10–100 CFU/ml) with any of the following organisms:

  • Staphylococcus aureus

  • Pseudomonas aeruginosa

  • Clostridium sporogenes

  • Bacillus subtilis

  • Candida albicans

  • Aspergillus niger

8.4.5 Spiking shall be done by Microbiology In-Charge and kept confidential.
8.4.6 Perform sterility test by membrane filtration method.
8.4.7 Repeat for three consecutive days.

Acceptance Criteria:

  • Unspiked samples: Must pass sterility test.

  • Spiked samples: Must fail sterility test.

Issue qualification certificate upon successful completion.

8.5 Microbial Limit Test Qualification

8.5.1 Perform MLT as per SOP.
8.5.2 Conduct testing for three consecutive days.

Acceptance Criteria:

  • %RSD should not exceed 1.0%.

Record observations and issue qualification certificate upon success.

8.6 Bioburden Test Qualification

8.6.1 Perform test as per Bioburden SOP.
8.6.2 Conduct testing for three consecutive days.

Acceptance Criteria:

  • Bacterial Count: NMT 10 CFU/100 ml

  • Fungal Growth: Absent

  • %RSD: ≤ 1.0%

Record results and issue qualification certificate.

8.7 Requalification Criteria

  • Every new analyst at the time of joining.

  • Annual requalification (once every year).

  • Requalification required after prolonged absence or repeated OOS linked to analyst error.

9.0 ANNEXURES

Annexure I – Test Report of Analyst Qualification for BET
Annexure II – Test Report of Analyst Qualification for Particulate Matter
Annexure III – Preparation & Spiking Record
Annexure IV – Test Report for Sterility Qualification
Annexure V – Test Report for %RSD Qualification
Annexure VI – Analyst Qualification Certificate
Annexure VII – Test Report for Bioburden Qualification

ANNEXURE – I

REPORT OF ANALYST QUALIFICATION FOR BACTERIAL ENDOTOXIN TEST (BET)

1. Analyst Information

Name of Analyst : ___________________________________________

Designation : _______________________________________________

Report No. : ________________________________________________

Trial No. : _________________________________________________

Date of Testing : ___________________________________________

2. Reagent Information

Parameter LAL Reagent Control Standard Endotoxin (CSE) LAL Reagent Water (LRW)
Lot No. __________ __________ __________
Expiry Date __________ __________ __________
Date of Reconstitution __________ __________ __________
Valid Up To __________ __________ __________
Sensitivity (λ) __________ N/A N/A

3. CSE Dilution Preparation

Tube No. CSE Volume (ml) LRW Volume (ml) Final Endotoxin Concentration (EU/ml)
1 ______ ______ ______
2 ______ ______ ______
3 ______ ______ ______
4 ______ ______ ______
5 ______ ______ ______
6 ______ ______ ______

4. Observations

Tube No. Endotoxin Concentration 2λ λ 0.5λ 0.25λ NWC End Point Log End Point
1 ______ ___ ___ ___ ___ ___ ___ ___
2 ______ ___ ___ ___ ___ ___ ___ ___
3 ______ ___ ___ ___ ___ ___ ___ ___
4 ______ ___ ___ ___ ___ ___ ___ ___

Σe (Sum of Log of End Points) = ______________________

5. Incubation Details

Temperature : ______________________

Heating Block ID No. : ______________________

Incubation Time :
From __________ To __________

6. Interpretation of Results

  • “–Ve” indicates No Gel Formation

  • “+Ve” indicates Gel Formation

7. Calculation

Geometric Mean (GM) Endpoint Concentration

GM = Antilog (Σ Log of End Points / Number of Replicates)

Calculated GM Endpoint Concentration = __________________ EU/ml

8. Final Result

Analyst Qualification / Re-Qualification Status:

☐ Complies
☐ Does Not Comply

As per applicable pharmacopoeial requirement (IP/BP/USP/In-house specification).

Done By Checked By
Name: __________________ Name: __________________
Signature: _____________ Signature: _____________
Date: __________________ Date: __________________

 

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