ELISA for evaluating the incorporation of plasma derived complement split-products C3b/iC3b into solid-phase immune complexes.
An ELISA that measures plasma derived complement (C) split-products C3b/iC3b deposited on solid-phase immune complexes during C activation is described. Plates are coated with BSA, anti-BSA and plasma is added. Deposited C3b/iC3b is then detected by biotinylated anti-C3c-antibodies, avidin-alkaline phosphatase and para-nitrophenylphosphate. A novel feature is that the assay measures residual C activation capacity rather than in vivo generated C activation products. The assay was applied to plasma from 250 healthy blood donors. No difference in activation capacity of either the alternative (AP) or classical pathway (CP) with regard to age or gender was demonstrated. The total coefficient of variation was <5.7%. The ELISA procedure was compared to a standard hemolytic complement CH(50) assay using plasma from 23 out-patients with systemic lupus erythematosus (SLE). There was a weak correlation between the two assays for both C pathways, but neither the ELISA nor the CH(50) assay showed any correlation with the diagnostic ACR-criteria for SLE. However, the capacity of the CP was significantly reduced in SLE out-patients compared to healthy blood donors (P<0.0001).
Department of Surgical Gastroenterology, Aalborg Hospital, Hobrovej 18-22, Postbox 365, 9100, Aalborg, Denmark. otu@aas.nja.dk
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Zimmermann-Nielsen
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Svehag
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Thorlacius-Ussing
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Baatrup
D000936:Antigen-Antibody Complex / Q000276:immunology; D003179:Complement C3b / Q000276:immunology*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D006801:Humans; D012680:Sensitivity and Specificity
D000936:Antigen-Antibody Complex; D003179:Complement C3b