Testing of parenteral pharmaceuticals and medical devices for pyrogens (fever-inducing substances) is critical to patient safety. The original rabbit pyrogen test has largely been replaced by different bacterial endotoxin tests based on Limulus amebocyte lysate (LAL), sourced from the blood equivalent of horseshoe crabs after comparative studies to the rabbit pyrogen test. Since 2004 a bacterial endotoxin test based on recombinant factor C (rFC), the endotoxin sensor protein inside of LAL, has been used as an animal-free alternative to LAL. Likewise, numerous studies compared LAL and rFC. Here we describe the history of pyrogen and bacterial endotoxin testing and summarize the evidence presented by those studies. We demonstrate that rFC and LAL are equivalent and comparable.
Some users have been hesitant to take on the validation of alternative methods. However, recombinant factor C (rFC) tests are so highly analogous to the horseshoe crab-sourced Limulus amebocyte lysate (LAL) tests that users should consider their advantages. They include sustainability, safeguarding the supply chain due to geographical diversity, accuracy, specificity, and reproducibility. Also noteworthy is that rFC is gaining acceptance in most of the world, with its own compendial chapter in the European pharmacopoeia (2.6.32)(1) and a new section written in the Chinese Pharmacopeia. Although given the delay of the USP, let us consider the "ease of validation" according to USP <1225> (2) as detailed in Q&A format below.What needs to be done for rFC Method Validation? With an operationally qualified reader installed, generally a full validation can be achieved in as little as 2 days, with 6 rFC assays, incl. 2 operators and 3 reagent lots to determine all necessary validation parameters.
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