Winter 2012 Volume 16 Number 1 1 PEER REVIEW: MICROBIOLOGY INTRODUCTION The pharmacopeial standard applying to sterile products is that sterile products must be capable of passing a test for sterility. The sterility test is an important, microbiological test in terms of regula- tory expectation rather than probability of detecting gross contamination. It allows for the examination of products purportedly to be sterile. This paper fo- cuses on the conventional sterility test method as set out in the harmonized pharmacopoeias (i.e., Euro- pean, Japanese, and United States). The sterility test is used as a product release test, where the sterility of a product is defined by the absence of viable and ac- tively multiplying microorganisms when the product or pharmaceutical preparation is tested in specified culture media. Detectable microbial growth is con- firmation of non-sterility, whereas sterility can be as- sumed from the absence of growth. In other words, the test assumes sterility unless non-sterility can be demonstrated. A failure with product sterility could lead to an adulterated product should the product be released onto the market (1). The method for conducting the sterility test is clearly documented in the European, Japanese, and United States pharmacopeias. Oc- casionally, the sterility test will produce a positive result. This demands both an examination of the laboratory test and an examination of the production process to determine why the sterility test failure occurred. The conclusion of such an investigation will be either that the sterility test was invalid due to some type of “laboratory error” (a position for which a great deal of caution is required given that regula- tory agencies require a robust rationale) or that the product was contaminated due to some event or inci- dent in the manufacturing, sterilization, or filling of the product or with the sterilization of components. This paper examines some of the areas to consider when looking at sterility test failures and in doing so draws upon the guidance in the FDA Guideline on Sterile Drug Products Produced by Aseptic Processing and from the author’s experiences of failure investi- gations (2). The emphasis is on the established con- ventional method. Although it is acknowledged that some companies have adopted rapid methods and in this context some of the guidance in this paper is not applicable, the conventional sterility test remains the most widely practiced and one of the most important release tests for sterile products. STERILITY TESTING The conventional sterility test is one of the oldest and most established pharmaceutical microbiology tests (3). It was first described, as a direct transfer meth- od into broth media, in the British Pharmacopeia in This paper examines the harmonized compendial ste- rility test (established by the United States, European, and Japanese pharmacopeias) and presents the key considerations for investigating sterility test failures. The emphasis is upon the conventional sterility test method. The paper focuses on the likely occurrences of false positives and the areas for consideration including the sterility test environment, operator technique, consumables, and reagents. This paper includes reference to genotypic microbiological identi- fication and current US Food and Drug Administration recommendations in relation to the number of permit- ted repeat sterility tests. Sterility Test Failure Investigations Tim Sandle