Capillary electrophoresis-based analysis of phospholipid and glycosaminoglycan binding by human β2-glycoprotein

Human beta(2)-glycoprotein I (beta(2)gpI) is a phospholipid and heparin binding plasma glycoprotein involved in autoimmune diseases characterized by blood clotting disturbances (thrombosis) together with the occurrence of autoantibodies against beta(2)gpI. With the final goal of assessing autoantibody influence on binding interactions of beta(2)gpI we have studied the development of capillary electrophoresis (CE)-based assays for interactions of negatively charged ligands with beta(2)gpI. In the development of suitable conditions for analysis at neutral pH of this basic protein (pI about 8) we found the pH hysteresis behavior of fused silica surfaces useful since the protonated surface after an acid pre-wash counteracted protein adsorption efficiently in contrast to more laborious procedures including acrylamide/dimethylacrylamide coatings that did not permit analysis of this particular protein. This simple approach made estimates of heparin-beta(2)gpI interactions possible and the principle was shown also to work for detection of beta(2)gpI binding to anionic phospholipids. Utilizing the pH hysteresis effect may be a simple solution to the adsorption problems often encountered in analyses of proteins by CE. (C) 2004 Elsevier B.V. All rights reserved.