Generation of anti-human apolipoprotein E isoform-specific monoclonal antibodies.

Abstract

The objectives of the present study were: (i) to establish an immunization protocol for the preparation of monoclonal antibodies (mAbs) specific for epitopes that are polymorphic in humans, (ii) to use this protocol to prepare anti-apoE mAbs that are either apoE isoform-specific or are antibody mimetics of the lipoprotein (LDL) receptor. We immunized mice that carry a transgene encoding apoE of one isoform with a second isoform of human apoE. Two immunization protocols were used. In protocol 1, mice that carried an apoE3 transgene were immunized with either apoE4), a natural apoE variant apoE that is defective in binding to the LDL receptor, or both. We hoped to obtain anti-apoE4-specific mAbs. In protocol 2, apoE transgenic mice were immunized with apoE3 and apoE4. We anticipated that we would generate mAbs that recognize apoE3 but not apoE4 and mAbs that recognize the same conformational apoE epitope that is recognized by the LDL receptor. Mice showed a weak anti-apoE immune response that had the predicted isoform specificity. Only one of the nine fusions generated stable, apoE-specific hybridomas. Of the 7 anti-apoE mAbs obtained, none possessed the isoform-specificity predicted from protocol 1. Two of the mAbs, 9F3 and 7D4 reacted well with apoE3 and apoE4 but failed to react with apoE2. The epitopes for both mAbs were mapped to a region that coincides with the apoE LDL receptor-binding site. Other anti-apoE mAbs could effectively compete with 7D4 and 9F3 for binding to immobilized apoE. (Abstract shortened by UMI.)