Studies on immunological assay of vitamin K dependent factors. II. Comparison of four immunoassay methods with functional activity of protein C in human plasma.

We describe and compare five assay systems for Protein C (PC) in human plasma; a functional assay for PC activity, Laurell electroimmunoassay with EDTA or calcium (EDTA-Laurell or Ca-Laurell), radioimmunoassay (RIA) and immunoradiometric assay (IRMA). The lower limit of sensitivity of PC in normal reference plasma was 2 X 10(-3) units/ml by RIA and 1 X 10(-4) units/ml by IRMA, but the ratio of maximum binding to blank binding was superior in RIA. In normal plasma (n = 20), there was no significant discrepancy of PC levels between RIA, Laurell (EDTA- and Ca-) and functional assay. In normal serum, PC antigen (PC:AG) measured by Ca-Laurell showed a high value (1.37 +/- 0.24 units/ml) compared to the other assays. PC:AG of a purified sample increased its level by Ca-Laurell method during activation by thrombin. Activated PC (PCa) has decreased affinity for anti-PC rabbit IgG, similarly to decarboxy (warfarinized) PC. In warfarin-treated individuals (n = 12) who were anticoagulated orally for more than 3 weeks, functional activity was lower (0.27 +/- 0.12 units/ml) than that measured by RIA (0.64 +/- 0.12 units/ml) or EDTA-Laurell (0.62 +/- 0.06 units/ml), whereas PC:AG measured by CA-Laurell had a normal value of 0.96 +/- 0.40 units/ml. After stopping administration, the activity showed a gradual increase to the normal at 2 weeks, while PC:AG increased more rapidly to normalize in 1 week.