The interaction between factor VIII clotting antigen (VIIICAg) and phospholipid

Abstract

The interaction between factor VIII clotting antigen (VIIICAg) and phospholipid (PL) was studied using a two-site solid phase immunoradiometric assay (IRMA) for VIII CAg. Incubation (2 h, 37 degrees C) of normal plasma, cryoprecipitate or factor VIII concentrate with Diagen PL (0.5 mg/unit VIIICAg) resulted in 80-90% loss of IRMA-measurable VIIICAg. No loss of factor VIII related antigen (VIIIRAg) or factor VIII clotting activity (VIIIC) was seen. Treatment of factor VIII concentrate with purified PLs showed greatest VIIICAg reduction with phosphatidylserine, less with phosphatidylethanolamine, and very little with phosphatidylcholine. The action of phospholipase-C (PL-C) on VIIICAg-PL complexes was investigated, with enzyme activity being monitored by thin-layer chromatography. Treatment of normal plasma, cryoprecipitate or factor VIII concentrate with PL-C (5 u/unit VIIICAg) resulted in 25%, 25% and 30% increases in VIIICAg. No increase in VIIIC or VIIIRAg was seen. Preincubation of factor VIII concentrate with PL, followed by PL-C incubation, resulted in 70-80% recovery of measurable VIIICAg. Incubation of 'activated' prothrombin complex with PL-C increased VIIICAg by 44% (Autoplex) and 80% (FEIBA), indicating VIIICAg-PL complexes are present. Incubation of factor VIII concentrate with fresh platelet lysate led to a reduction in VIIICAg (100 u/dl to 21 u/dl). In fresh platelet lysate alone low VIIICAg levels were detectable (0.71 X 10(-3) u/10(9) plt). After PL-C incubation VIIICAg levels increased to 9.8 X 10(-3) u/10(9) plt) (14-fold increase). Thus VIIICAg in platelets may be hidden by VIIICAg-PL complexes.