Evaluation of growth based rapid microbiological methods for sterility testing of vaccines and other biological products

Abstract

Most biological products, including vaccines, administered by the parenteral route are required to be tested for sterility at the final container and also at various stages during manufacture. The sterility testing method described in the Code of Federal Regulations (21 CFR 610.12) and the United States Pharmacopoeia (USP, Chapter ) is based on the observation of turbidity in liquid culture media due to growth of potential contaminants. We evaluated rapid microbiological methods (RMM) based on detection of growth 1) by adenosine triphosphate (ATP) bioluminescence technology (Rapid Milliflex(®) Detection System [RMDS]), and 2) by CO(2) monitoring technologies (BacT/Alert and the BACTEC systems), as alternate sterility methods. Microorganisms representing Gram negative, Gram positive, aerobic, anaerobic, spore forming, slow growing bacteria, yeast, and fungi were prepared in aliquots of Fluid A or a biological matrix (including inactivated influenza vaccines) to contain approximately 0.1, 1, 10 and 100 colony forming units (CFU) in an inoculum of 10 ml. These preparations were inoculated to the specific media required for the various methods: 1) fluid thioglycollate medium (FTM) and tryptic soy broth (TSB) of the compendial sterility method (both membrane filtration and direct inoculation); 2) tryptic soy agar (TSA), Sabouraud dextrose agar (SDA) and Schaedler blood agar (SBA) of the RMDS; 3) iAST and iNST media of the BacT/Alert system and 4) Standard 10 Aerobic/F and Standard…