SOP on Viable Particulate Monitoring – Sterility Testing Area
1.0 Objective
1.1 To lay down the procedure for Viable Particulate Monitoring in the Sterility Testing Area of the Microbiology Laboratory to ensure compliance with acceptable limits of total viable count.
2.0 Scope
2.1 This SOP is applicable to viable particulate (microbial) monitoring of the Sterility Testing Area in the Microbiology Laboratory.
3.0 Responsibility
3.1 Chemist or above – Microbiology Laboratory: Execution of environmental and personnel monitoring activities and documentation.
3.2 Head – Microbiology Section: Review of monitoring results and ensuring compliance.
4.0 Accountability
4.1 Head – Quality Control: Overall implementation and compliance.
4.2 Head – Quality Assurance: Oversight and review of compliance with environmental monitoring requirements.
5.0 Procedure
5.1 General
5.1.1 Viable particulate monitoring in the sterility testing facility shall be performed to ensure that environmental conditions comply with specified microbiological limits.
5.1.2 Monitoring shall be performed using the following methods:
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Settle Plates (Passive Air Sampling)
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Volumetric Air Sampling (Active Air Sampling – Sieve Impaction Method)
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Surface Monitoring by Swabs and 55 mm Surface Contact (RODAC) Plates
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Personnel Monitoring by 55 mm Surface Contact (RODAC) Plates
5.2 Preparation of Media and Accessories
5.2.1 Prepare and sterilize required SCDA (Soyabean Casein Digest Agar) plates/cassettes and pre-incubate as per SOP (Sterile Media Preparation).
5.2.2 Sterilize required tubes containing 10 ml sterile saline and sterile swabs. Arrange in test tube stands.
5.2.3 Sterilize required stainless steel (SS) carriers for transporting plates and accessories.
5.2.4 Place pre-incubated media plates/cassettes under LAF working zone.
5.2.5 Disinfect outer surfaces of media plates/cassettes using 1% Protasan DS / 1% Combatan / 1% Triple 100 and place them in sterile SS carriers along with swab stands.
5.2.6 Transfer carriers to sterilization area.
5.2.7 Disinfect outer surfaces of carriers and keep them in dynamic pass box (cooling zone) under UV light for 30 minutes.
5.2.8 Enter sterility testing area through change rooms as per SOP .
5.2.9 Ensure air sampler is fully charged.
5.2.10 Disinfect outer surface of SS carrier and bring inside the sterility area.
5.3 Settle Plate Method (Passive Air Sampling)
5.3.1 Expose SCDA plates at predefined locations as per annexure schematic diagrams.
Exposure Procedure:
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Bring plate to sampling location.
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Carefully open the lid.
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Place plate on exposure stand (if provided).
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Keep lid inverted beside plate (inner side down).
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Expose for 4 hours.
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Close plate after exposure.
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Label bottom of plate with location code, exposure date, media lot number, and media preparation date.
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Place exposed plates in SS carriers.
5.3.2 Frequency:
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Daily once under dynamic conditions (if activity present).
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Once per week if no activity.
5.4 Volumetric Air Sampling (Active Air Sampling)
5.4.1 Perform sampling at predefined locations (1000 liters per location) as per Annexure.
5.4.2 Operate sieve impaction air sampler as per SOP.
5.4.3 Disinfect sieve after sampling at each location.
5.4.4 Perform sampling sequentially from highest clean area to lowest clean area.
5.4.5 Label plates with location code, exposure date, media lot number, and preparation date.
5.4.6 Frequency: Once per week.
5.5 Surface Monitoring
5.5.1 Swab Method
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Aseptically open sterile swab tube.
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Remove excess saline by pressing against inner wall.
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Swab approximately 25 cm² (5 cm × 5 cm) area using horizontal strokes followed by vertical strokes (after 90° rotation).
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Return swab to saline tube and close.
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Label appropriately.
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Place swab tubes in SS carriers.
5.5.2 RODAC Plate Method (55 mm Surface Contact Plates)
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Carefully open RODAC plate.
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Invert and gently press convex agar surface onto flat surface.
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Ensure full agar contact.
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Remove and close lid carefully.
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Disinfect sampled surface.
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Label plate appropriately.
5.5.3 Frequency:
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Daily at end of sterility testing (before cleaning).
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Fortnightly if no activity.
5.6 Personnel Monitoring
5.6.1 Perform monitoring using 55 mm RODAC plates.
5.6.2 Monitor four mandatory locations including both hands, plus two additional locations as per Annexure .
5.6.3 Procedure:
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Open RODAC plate carefully.
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Press agar surface onto sampling area.
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Close lid and label appropriately.
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Place plates in SS carriers.
5.6.4 Frequency: Daily at end of activity before exiting sterility area.
5.7 Post-Monitoring Handling
5.7.1 Place SS carriers in dynamic pass box and exit through change rooms.
5.7.2 Transfer all plates, cassettes, and swab tubes to incubator room.
5.7.3 For swabs:
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Vortex saline tube with swab.
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Filter through sterile 0.45 µm membrane filter.
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Rinse filter three times with 100 ml of 0.1% peptone water.
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Place membrane filter on pre-incubated SCDA plate.
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Use separate filter holder for each swab.
5.8 Incubation
5.8.1 Incubate all environmental and personnel monitoring plates/cassettes:
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20–25°C for 72 hours
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Followed by 30–35°C for additional 48 hours
5.8.2 Observe and record results after 72 hours and after completion of 48 hours.
5.9 Documentation
5.9.1 Record Settle Plate results as per Annexure.
5.9.2 Record Volumetric Air Sampling results as per Annexure.
5.9.3 Record Surface Monitoring results as per Annexure.
5.9.4 Record Personnel Monitoring results as per Annexure.
5.10 Acceptable Levels
5.10.1 Verify that monitoring results comply with acceptable microbiological limits specified in respective annexures.
5.10.2 Any result exceeding limits shall be investigated as per deviation/OOS procedure.
6.0 List of Annexures
Annexure – I: Report of Viable Particles Monitoring by Settle Plate Method
Annexure – II: Report of Viable Particles Monitoring by Volumetric Air Sampling
Annexure – III: Report of Surface Monitoring by Swabs and RODAC Plates
Annexure – IV: Report of Personnel Monitoring by Surface Contact Plates
SOP For Handling of Microbiological Data Deviation (MDD) in Microbiology Laboratory