Differential blocking of coagulation-activating pathways of Limulus amebocyte lysate
| Autor: | Gui-Hang Zhang, Claus Koch, Ole Buchardt, L. Baek |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Cell Extracts
Lipopolysaccharides Microbiology (medical) beta-Glucans Lipopolysaccharide Enzyme-Linked Immunosorbent Assay Curdlan Biology Sensitivity and Specificity Arthropod Proteins Microbiology Mice chemistry.chemical_compound Laminarin Polysaccharides Hemolymph Horseshoe Crabs medicine Animals Dimethyl Sulfoxide False Positive Reactions Glucans Limulus Test Polymyxin B Enzyme Precursors Blood Cells Alcaligenes faecalis Dimethyl sulfoxide Serine Endopeptidases Antibodies Monoclonal biology.organism_classification Molecular biology Endotoxins Enzyme Activation chemistry Limulus amebocyte lysate Limulus Research Article medicine.drug |
| Zdroj: | Journal of Clinical Microbiology. 32:1537-1541 |
| ISSN: | 1098-660X 0095-1137 |
| Popis: | The coagulation of Limulus amebocyte lysate (LAL) can be activated through two pathways, one initiated by endotoxin and the other by beta-glucans. The two pathways join at the step of activation of the proclotting enzyme. We report here that the endotoxin-activated pathway can be differentially inhibited by two methods in a Limulus enzyme-linked immunosorbent assay (ELISA), either by the combined use of dimethyl sulfoxide and polymyxin B or by a monoclonal antibody against Limulus factor C. LAL reactivities to 10 different endotoxin preparations could be inhibited by the former method by a factor of 10(4) to 10(6) and could be blocked almost totally by the latter method, irrespective of the source of endotoxin. The sensitivity of the assay was approximately 50 pg/ml both for curdlan from Alcaligenes faecalis and for laminarin from Laminaria digitata. We also found that the beta-glucan-activated pathway could be totally blocked by laminarin (> 1 microgram/ml) without affecting the endotoxin-activated pathway, allowing endotoxin to be quantitated specifically by the Limulus ELISA with a detection limit of 0.005 endotoxin unit per ml. The use of uninhibited and differentially inhibited ELISAs demonstrated that different LAL preparations showed much greater variation in assaying beta-glucans than in assaying endotoxins. The LAL reactivity of normal human plasma was found to be due to the activation of the beta-glucan pathway, but not the endotoxin pathway, of LAL. |
| Databáze: | OpenAIRE |
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