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The European Pharmacopoeia (Ph. Eur.) provides principles for microbiological testing of tissue preparations. According to the Ph. Eur., tests should be performed at different temperatures for detection of aerobic bacteria and fungi (20–25°C) vs. anaerobic bacteria (30–35°C). Semiautomated systems using blood culture bottles are already widely used and they are adequate for growth detection. Resin-containing bottles and the addition of penicillinase permit testing of culture media containing antibiotics.   Materials and Methods:   At 3 temperatures (21, 30, and 35°C) cornea culture media with and without dextran (CM II and CM I) and thermal disinfected femoral head medium (FH) were spiked with the 6 reference strains recommended by the Ph. Eur. (additionally:  Enterococcus faecalis ,  Staphylococcus epidermidis,  and  Cutibacterium acnes ). Microbial growth was monitored with the BACTEC<sup>TM</sup> FX unit or visually at 21°C.   Results:   Growth for all strains was detected with each medium at all 3 temperatures, except for  C. acnes  at 21°C (all media) and 30°C with FH.  C. acnes  had the highest times to detection, requiring test durations of 14 days. Microbial growth was faster at 30 and 35°C compared to 21°C.   Conclusion:   The requirements according to the Ph. Eur. for a successful method suitability test could be fulfilled for the semiautomated blood culture bottle system with the BACTEC<sup>TM</sup> FX unit for the media and microorganisms used. In the presented validation study 35°C was shown to be the incubation temperature with the fastest growth, of the majority of the test strains used, and complete detection within 14 days.

Validation of Microbiological Testing of Tissue Preparations with Different Incubation Temperatures