Validation Of Rapid Sterility Test Method For Cellular T

无菌检测是细胞治疗产品生产过程和产品放行检测的重要组成部分。FDA特别建议按照21 CFR 610.12进行无菌检测。此外，由于他们担心抗生素可能会干扰无菌测试的准确评估，FDA要求根据USP“<71>无菌测试”对所有含有抗生素的样品进行初步的抑菌和抑菌测试，包括在含有抗生素的培养基中生长的细胞。CFR和USP标准中描述的无菌检测方法是在25年前开发的，并且是劳动密集型的，需要培养14天。自从这些方法发表以来，已经开发出更灵敏和快速的方法来检测各种体液中的微生物生长。这些现代方法包括用于检测血液和其他通常无菌液体中微生物生长的自动化系统。尽管许多实验室使用这些自动化系统来评估细胞治疗产品的无菌性，但FDA尚未批准该申请，因为没有公布的数据表明这些新方法与CFR和USP方法（CFR/USP）进行了正式比较。由于这些原因，我们开发了一种验证方案，将CFR/USP方法与两种自动培养方法进行比较：生物梅里埃BacT/Alert （BTA）系统和Becton Dickinson （Bactec）系统。

Sterility testing is an essential part of in-process and release testing for cellular therapy products. The FDA specifically recommends that sterility testing be performed as outlined in 21 CFR 610.12. Furthermore, because they are concerned that antibiotics may interfere with the accurate assessment of sterility testing, the FDA requires preliminary bacteriostasis and fungistasis testing according to the USP "<71> Sterility Test" on all samples containing antibiotics, including cells grown in antibiotic-containing media. The methods for sterility testing described in the CFR and USP standards were developed more than 25 years ago, and are labor-intensive and require incubation for 14 days. Since the time these methods were published, more sensitive and rapid methods have been developed for detection of microbial growth in various body fluids. These modern methods include automated systems for the detection of microbial growth in blood and other normally sterile fluids. Despite the fact many laboratories use these automated systems to assess sterility of cellular therapy products, the FDA has not sanctioned this application because there are no published data from any formal comparison of these newer methods with the CFR and USP methods (CFR/USP). For these reasons, we have developed a validation protocol comparing the CFR/USP method with two automated culture methods: the bioMerieux BacT/Alert (BTA) system and the Becton Dickinson (Bactec) system. Six different cell products from the Department of Transfusion Medicine (DTM) were seeded with selected bacteria (8 strains) and fungi (2 strains) (10 colony forming units and 50 CFUs) and then tested in triplicate with each method. The test sensitivity and time to detect a positive culture were assessed. With each bacterium and fungus, the automated systems were found to be more rapid and sensitive for detection of contaminated cell products. Growth of strict aerobic and anaerobic organisms grew preferentially in aerobic and anaerobic blood culture broths, respectively; therefore, each cell product would have to be cultured in the two broth formulations. Results: Positive cultures were detected in a mean (range) of 72% (7-100%) of the cultures for CFR/USP, 82% (0-100%) for BTA, and 93% (57-100%) for Bactec. For nine of the 10 organisms tested, overall detection rates for BTA and Bactec were equivalent to or higher than CFR/USP. Of the six product media tested, detection of organisms was impaired only by the medium containing multiple antibiotics: this occurred in all three systems. Both BTA and Bactec had shorter times to detection than the CFR/USP method, with overall means (ranges) of 87 (24-264 ) hours for CFR/USP, 24 (12-54) hours for BTA, and 33 (12-80) hours for Bactec. Detection occurred coonsistently within 7 days for both BTA and Bactec, but not for CFR/USP.