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A modification of a two-site, solid-phase immunoradiometric assay (IRMA) for Factor VIII coagulant antigen (VIII: CAg) has been evaluated for its potential to detect antibodies against Factor VIII coagulant activity (VIII: C) in patient plasma samples. For this purpose the assay system comprised four steps: 1) coating of test tubes with human anti —VIII:C , 2) incubation with normal Factor VIII— VWF complex, 3) incubation with test sample, and 4) binding of radiolabeled human anti–VIII: C as marker.  Of eight hemophilic plasma samples containing antibodies against VIII: C (as detected in a clotting assay) five were able to prevent binding of radiolabel partially and three prevented this completely. One of these three (which actually was the antibody used in the IRMA), was effective even at very high dilutions. Two hemophilic plasma samples without detectable antibodies in the clotting assay, a severe Von Willebrand’s disease plasma and normal plasma samples showed no significant interference with binding of radiolabeled human anti-VIII: C. Also, a plasma sample containing a high titer of spontaneous human antibody to VIII: C as well as a heterologous antiserum against Factor VIII—VWF complex did not interfere with binding of radio-activity.  It is concluded that the test system described is a sensitive tool to detect antibodies of the same specificity as those used in the IRMA. It may also detect antibodies of differing specificity. The lack of crossreactivity with some antibodies points to interindividual differences in specificity of antibodies against VIII: C.

Immunoradiometric Assay Of VIII:CAG, A Potential Tool To Detect Human Anti-VIII:C Antibodies