Simple immunochromometric assay for protein C activity

Abstract

Protein C binds readily from human plasma to antibody-coated wells, where it may be quantitated with an iodine 125-labeled antibody to protein C. Treatment with thrombin results in a small reduction in the protein C antigen detectable by this immunoradiometric assay (IRMA). However, activated protein C resulting from thrombin treatment and retained by the antibody on a solid phase may be detected by an overnight incubation with chromogenic substrates S-2366 or CBS 34.47. The immunochromometric assay (ICMA, analogous to IRMA) described uses a heterologous antibody to protein C, activation with relatively low concentrations of bovine thrombin, and quantitation by hydrolysis of chromogenic substrate in a convenient 96-well microtiter plate system. The correlation between IRMA and ICMA protein C results was found to be good with normal persons and patients with liver disease. Patients taking oral anticoagulants had reduced protein C antigen (IRMA) but even lower protein C by ICMA, indicating that inactive forms were present.