[ HOW-TO GUIDE ]
How to Investigate a Sterility Test Positive
Decide whether a sterility test failure is a laboratory artefact or a real sterility failure — and act on either.
What a how-to is not
A how-to is SPEQ’s practitioner method, not a procedure. It does not replace your own SOP, it is not a validated approach, and the judgement calls in it belong to your quality unit.
A positive sterility test is a specification failure, and 21 CFR 211.192 requires it to be thoroughly investigated, extended to associated batches and products, and recorded with conclusions and follow-up. EU GMP Annex 1 (2022) adds that the sterility test is only the last of a series of controls, so the investigation has to examine those controls, not only the laboratory.
- 1
Secure the evidence
Quarantine the batch, retain the positive media and the isolate, and preserve the laboratory records, the test environment monitoring data and the sample handling records before anything is cleaned or discarded.
- 2
Identify the organism
Identify the isolate to species and compare it with the laboratory environment, the manufacturing area’s environmental and personnel monitoring flora, and the bioburden data for the batch. The identity is the strongest single indicator of where it came from.
- 3
Examine the laboratory
Review the test performance: aseptic technique, the isolator or cleanroom used, negative controls, media quality control, and any process used to decontaminate sample surfaces, which Annex 1 says must not affect the test’s sensitivity or the sample’s reliability.
- 4
Examine manufacturing
Review the batch against its sterility assurance: sterilisation or filtration records, filter integrity results, environmental and personnel monitoring for the filling session, interventions, aseptic process simulation history and, for terminally sterilised product, load mapping and cycle data.
- 5
Extend and conclude
Extend the investigation to other batches and products that may be associated with the failure, as 21 CFR 211.192 requires. Record the conclusion, the evidence for it, the batch disposition and the CAPA. Conclude laboratory error only when the evidence supports it, not by default.
- !Invalidating the test as a laboratory error without evidence, then releasing on a retest.
- !Not comparing the isolate with the manufacturing area’s monitoring flora.
- !Limiting the investigation to the one batch, when 211.192 requires extension to associated batches and products.
- !Forgetting that a passing sterility test cannot assure sterility of a batch that did not meet its process parameters.
How to Investigate a Sterility Test Positive: frequently asked questions
Common questions on investigate a sterility test positive.
Does US GMP require the investigation to look beyond the failing batch?
Yes. 21 CFR 211.192 says the investigation of a batch failure shall extend to other batches of the same drug product and other drug products that may have been associated with the specific failure, with a written record including the conclusions and follow-up.
Can a passing sterility test compensate for a process that went outside its validated parameters?
No. Annex 1 (2022) paragraph 10.5 says the sterility test cannot be used to assure sterility of a product that does not meet its design, procedural or validation parameters; it is the last in a series of critical control measures.
Which samples should be tested for sterility from an aseptic fill?
Annex 1 paragraph 10.6 says samples should represent the whole batch and include containers filled at the beginning and end of the batch, with additional samples, for example after critical interventions, considered based on risk.
WHERE NEXT
- DO ITOpen the Out-of-Specification (OOS) Investigation Report template →The structure these steps fill in.
- UNDERSTAND ITSterility Testing →The principles behind the procedure.
- ALL GUIDESBrowse all 127 how-to guides →
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