A new sensitive microplate assay of plasma endotoxin

✍ Scribed by Dr. Hiroshi Tamura; Shigenori Tanaka; Taminori Obayashi; Minoru Yoshida; Tadashi Kawai

• Publisher: John Wiley and Sons
• Year: 1992
• Tongue: English
• Weight: 387 KB
• Volume: 6
• Category: Article
• ISSN: 0887-8013
• DOI: 10.1002/jcla.1860060412

✦ Synopsis

We have developed a microplate method for determining endotoxin in platelet-rich plasma-using Endospecy, an endotoxinspecific chromogenic Limulus test reagent. Nonspecific activators and inhibitors of the test were eliminated by exposing samples (5 FI) to the alkali reagent consisting of KOH, CaCI,, Triton X-1 00, ethyleniminepolymer and N,N-bis(2-hydroxyethyl)glycine. The recoveries of various endotoxins were almost complete and not enhanced by dilution. The doseresponse curve was linear over endotoxin concentrations of 2-400 pg/ml with good precision (C.V. -4.0%). Normal human plasmas

Key words:

(n =30) contained -4 . 0 pg/ml of endotoxin in reference to that of Escherichia coliolll: 84. All plasma samples with high concentration of endotoxin by a conventional method showed high values by the microplate assay as well. Since it does not require centrifugation, the new treatment allows the whole reactions to proceed on the same microplate. This permits us to apply the Limulus test to an automated assay system, making plasma endotoxin determination simpler and more rapid than a conventional test tube method.

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Limulus test, amebocyte lysate, chromogenic substrate, platelet-rich plasma (PRP), factor G