Changes in high molecular weight kininogen levels during and after cardiopulmonary bypass surgery measured using a chromogenic peptide substrate assay.

Abstract

High molecular weight kininogen (HK) is a co-factor in the blood-contact activation system. A chromogenic peptide substrate assay for HK (HKcs) has been developed in which test plasmas are mixed with diluted HK-deficient plasma and incubated with a soluble contact system activator that activates prekallikrein and factor XII. Calcium chloride, a synthetic thrombin inhibitor and a chromogenic peptide substrate for activated factor X (FXa) are then added. The FXa generated cleaves the FXa substrate releasing p-nitroanaline, which is measured photometrically. Test plasma HK values were calculated from a standard curve generated using a pooled normal plasma. Acceptable intra-assay and inter-assay precision values were obtained and levels of HK up to 200% were measurable. The assay measured HK in plasmas deficient in factor XII, prekallikrein and factor XI, was not affected by antiphospholipid antibodies and gave an acceptable correlation (r = 0.95) when normal plasmas and mixtures of HK-deficient and normal pooled plasma, calculated to give HK levels of 25 and 50%, were compared using HKcs and a HK one-stage clotting assay. The HKcs was used to measure HK levels in seven patients undergoing cardiopulmonary bypass (CPB). HK levels fell significantly during CPB (P = 0.0014) and were significantly higher (P = 0.016) 6 days after CPB, suggesting that HK may be a positive acute-phase reacting protein.