Mycoplasmas are potential contaminants that introduce undesirable changes in mammalian cell cultures. They frequently contaminate cell substrates and other starting materials used for manufacturing cell-derived biologics, such as vaccines and pharmaceutical products. Mycoplasma purity testing of live vaccines, active ingredients, raw material, and seed lots is required during vaccine production. Previously, testing using a time-consuming, costly 28-day culture assay, which lacks sensitivity for species that do not grow in culture, was required in the European Pharmacopoeia (Ph. Eur). But now nucleic acid amplification techniques (NATs) can be used. NATs provide rapid results and are sensitive. We evaluated the sensitivity and specificity of a commercially-available NAT to detect individual mycoplasma DNA in a veterinary modified live vaccine using five reference strains recommended by the Ph. Eur. Our results showed that this NAT-based method can be used to detect mycoplasma in spiked live vaccine, without interference from the vaccine components, with a limit of detection of 10 CFU/mL, as required by the Ph. Eur. Its specificity was demonstrated since no mycoplasmas were detected in non-spiked vaccine. This method is undergoing validation as a replacement for the conventional culture method in the production of veterinary live vaccines.
Keywords: DNA chips; Limit of detection; Mycoplasma; PCR; Vaccine.
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