Open AccessA new sensitive microplate assay of plasma endotoxin
Author(s) -
Tamura Hiroshi,
Tanaka Shigenori,
Obayashi Taminori,
Yoshida Minoru,
Kawai Tadashi
Publication year - 1992
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.1860060412
Subject(s) - chromogenic , chromatography , chemistry , reagent , centrifugation , limulus , dilution , biology , paleontology , physics , thermodynamics
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 (5μl) to the alkali reagent consisting of KOH, CaCl 2 , Triton X‐100, ethyleniminepolymer and N,N‐bis(2‐hydroxyethyl)glycine. The recoveries of various endotoxins were almost complete and not enhanced by dilution. The dose—response curve was linear over endotoxin concentrations of 2–400 pg/ml with good precision (C.V.<5.0%). Normal human plasmas (n=30) contained <5.0 pg/ml of endotoxin in reference to that of Escherichia coli 0111:B4. 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. © 1992 Wiley‐Liss, Inc.