Bacterial endotoxin testing
Kinetic chromogenic, turbidimetric and gel-clot LAL; why the index specification is a screening threshold and not a limit; and the dose arithmetic that gives a real limit.
Microbiological
Sterility testing
Membrane filtration and direct inoculation to USP <71>: the media, the fourteen days, and why a passed sterility test on one vial says very little.
A sterility test detects viable micro-organisms by incubating the article, or a filtrate of it, in growth media and observing for turbidity. It is a destructive test on a sample, it takes fourteen days, and its statistical power is poor by construction: testing twenty units from a lot of a thousand cannot detect a contamination rate below a few per cent with any confidence.
The index records sterility on the 9 % of reports where it was ordered, reports the method and the incubation period, and does not treat a pass as a statement about the lot.
Method
| Element | Requirement |
|---|---|
| Preferred technique | Membrane filtration, 0.45 µm, where the article is filterable |
| Alternative | Direct inoculation, where filtration is impracticable |
| Media | Fluid thioglycollate medium (anaerobes and aerobes) and soya-bean casein digest medium (fungi and aerobes) |
| Incubation | FTM at 30–35 °C, SCDM at 20–25 °C, not less than 14 days |
| Negative control | Media-only, incubated in parallel |
| Growth promotion | Each medium lot challenged with specified organisms and shown to support growth |
| Method suitability | The article must be shown not to inhibit growth of the challenge organisms at the concentration tested |
Method suitability is the step most often omitted. A peptide with antimicrobial activity — and LL-37 is precisely that, by design — will suppress growth in the test medium and produce a false pass. A sterility result on an antimicrobial peptide without a documented suitability test is not interpretable, and the index says so on those records.
What a pass and a fail mean here
- No growth at 14 d: no viable organisms were recovered from the units tested under these conditions. It is not a statement that the lot is sterile, and it is not a statement that any other vial is.
- Growth detected: an organism was recovered. This is a strong finding, and the index escalates it: the record is flagged, the organism is named where identified, and the batch page carries the result prominently. It remains possible that the contamination was introduced during testing, which is what the negative control exists to address.
- Not tested: the overwhelming majority. The index prints this explicitly rather than leaving the field blank, because a blank reads as a pass to most people.
Normative references
Cited because they are the documents the acceptance criteria above are taken from. The index applies them as written and states every deviation.
- USP <71> — Sterility tests
- Ph. Eur. 2.6.1 — Sterility
- USP <1211> — Sterility assurance
- USP <797> — Pharmaceutical compounding, sterile preparations
Related methods
Appearance, reconstitution clarity and pH
The three cheapest measurements on a vial, what each one actually diagnoses, and why the index prints them verbatim.
Physical tests