IMMUNOENZYMOMETRIC ANALYSIS FOR PLASMA VON-WILLEBRAND-FACTOR DEGRADATION IN DIABETES-MELLITUS USING MONOCLONAL-ANTIBODIES RECOGNIZING PROTEASE-SENSITIVE SITES

High-molecular-weight von Willebrand factor (vWf) multimers were separ ated from their smaller multimers by molecular-sieve chromatography an d were measured by several sandwich enzyme-linked immunosorbent assays (ELISA) with monoclonal antibodies (moABs) against human vWf. The epi topes of these moABs were mapped partially using fragments generated b y V-8 protease digests of native vWf. Large multimers were more immuno reactive with the sandwich ELISA using immobilized VW28-1 and enzyme-l abeled VW92-3 than with any other ELISA. The epitopes recognized by th ese two moABs were sensitive to trypsin and plasmin digestion, and the other moABs appeared to be reactive with the extensively digested ant igen. Relative reactivities of this ELISA to plasma vWf multimers in d iabetes mellitus were significantly reduced compared to those in healt hy subjects. These data demonstrate that molecular abnormalities of vW f circulating in diabetic patients may be caused by some plasma protea ses that increase in the microcirculation of patients with diabetic va scular diseases.