Quantitative comparison of in vitro and in vivo methods for the detection of endotoxin
Cooper, J.F.; Levin, J.; Wagner, H.N.
Journal of Laboratory and Clinical Medicine 78(1): 138-148
1971
ISSN/ISBN: 0022-2143 PMID: 4936365 Document Number: 5685
The administration of drugs or fluids which have become contaminated with bacterial endotoxin can produce shock, fever, and death. The current legally required bioassay for endotoxin in parenteral solutions is based on the pyrogenic effect of bacterial endotoxin in rabbits and is difficult to perform. The Limulus method for the detection of endotoxin is based upon the reaction in vitro between endotoxin and a lysate prepared from amebocytes, the circulating blood cells of Limulus. This assay is rapid, simple, and sensitive and requires only 0.1 ml. of the material being tested. The Limulus test detected 0.001 μg per milliliter of E. coli endotoxin, whereas injection of the same concentration according to the official protocol of the U. S. Pharmacopeia did not result in a pyrogenic response. Similarly, the Limulus test detected 0.0001 μg per milliliter of Klebsiella endotoxin, although a dose of 0.0006 μg per kilogram was not pyrogenic in rabbits. There was good correlation between the pyrogenic potency of purified endotoxins of E. coli and Klebsiella, as measured by their median-pyrogenic dose and fever index, and the rate of gelation of lysates of Limulus amebocytes. Two preparations of short-lived radiopharmaceutical agents which were pyrogenic produced gelation of amebocyte lysate in less than 15 minutes. The results demonstrated that the Limulus test was approximately 10 times as sensitive to endotoxin as the official pyrogen test. This in vitro assay is a potential alternative test for the detection of bacterial endotoxin in drugs, fluids, and other agents prepared for parenteral administration to human beings.
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