Standardization of an enzymometric assay for apolipoprotein A-I by using mixtures of monoclonal antibodies

Summary: For the Standardization of human plasma apolipoprotein A-I assay two well characterized mono-clonal antibody mixtures were used to develop a sandwich immunoenzymometric assay. The monoclonal antibody mixture l (A05-A17-A30) in solid phase technique was selected on the basis of its higher binding capacity of [125I]HDL (41 ng per well) compared to polyclonal antibody (23 ng per well). The epitopes recognized by monoclonal antibody mixture l are surface antigenic sites of apolipoprotein A-I expressed on native HDL äs determined by competitive imhibition of labeled HDL. The peroxidase conjugated monoclonal antibody mixture 2 (A03-A05-A17-A51) was selected on the basis of its ability to bind to apolipoprotein-captured by monoclonal antibody mixture 1. For this, we used the 125I-labeled monoclonal antibodies. Under optimized assay cönditioiis, the immunoenzymometric assay is precise (intra- and inter-assay coefficients of variations 5.4 % and 9.2 % respectively). It yields plasma apolipoprotein A-I values that correlate highly with those obtained with polyclonal antibody (r = 0.96). So the use of well characterized monoclonal antibody mixtures reacting only to surface antigenic sites of apolipoprotein A-I present on native lipoprotein may provide the possibility öf Standardization of apolipoprotein A-I measurement. n o uc on, ^ potential advantages of the enzymometric assa