Identification of HDL Binding Proteins by an Immunological Ligand Blotting Assay

Abstract

Recently, we identified two HDL binding proteins (Mr=120 K-Da and 100 K-Da) in rat and human liver plasma membranes by ligand blotting using ¹²⁵I-HDL₃, and purified these binding proteins. The detection method needed 3 or 4 days for its autoradiography.
Here, we report a method to visualize HDL binding proteins by ligand blotting using HDL₃, anti-apo A-I and HRPO-conjugated 2nd Ab. By this method, we identified the same binding proteins as those detected by ligand blotting using 125^I-HDL₃.
The advantages of this method are that we can visualize HDL binding proteins within 2 days and can use diluted serum as a ligand instead of radiolabeled purified HDL₃.