Development and characterization of stable reporter cells for fast and sensitive detection of pyrogen

摘要

Pyrogens are a class of heterogeneous compounds that cause fever and induce inflammatory responses in the host. Lipopolysaccharides (LPS, also known as endotoxin) is the major pyrogen in the category of drug quality control. Accurate and fast quantification of pyrogens is crucial for drug safety. In the present study, we aimed to develop a sensitive and reliable method for rapid detection of pyrogens using luciferase reporter assay. Stable human A549 luciferase reporter cells were constructed under the control of a NF-kappa B -responsive element or IFN-beta promoter. Our results showed that several monoclonal stable cell clones responded to 0.1 EU/ml endotoxin, which was less than human fever threshold at 0.3 EU/ml of endotoxin. Further, compared with original A549 cells, TLR4 expression on the reporter cells were significantly increased after low amount LPS stimulation. In addition, reporter cells also responded to zymosan stimulation. Therefore, these results indicated that the stable luciferase reporter cells respond to endotoxin and non-endotoxin pyrogens and have the potential to further develop into a sensitive and fast pyrogen evaluation method.