ELISA for Soluble Form of Lectin-Like Oxidized LDL Receptor-1, A Novel Marker of Acute Coronary Syndrome

Abstract

Lectin-like oxidized LDL receptor-1 (LOX-1) is prominently expressed in atherosclerotic plaques (1)(2). LOX-1 expressed on the cell surface is proteolytically cleaved at its membrane-proximal extracellular domain (ECD) and released in soluble form (sLOX-1) (2)(3). Serum sLOX-1 is increased at an earlier stage of acute coronary syndrome (ACS) than are other cardiac makers, such as creatine kinase-MB fraction, troponin T, and high-sensitivity C-reactive protein (4). We describe the development of an ELISA for sLOX-1. The cleavage sites on human LOX-1 (hLOX-1) remain unknown. We tried to obtain human sLOX-1 (hsLOX-1) from cultures of CHO-K1 cells stably expressing hLOX-1 (3)(5), but the amount of hsLOX-1 obtained was too small for ELISA development. hsLOX-1 should be similar to the ECD of hLOX-1 (hLOX-1-ECD), considering a structure of bovine sLOX-1 (3). We produced recombinant hLOX-1-ECD [rhLOX-1-ECD; hLOX-1 (84–273)] in Escherichia coli and purified it as described previously (5); we then used the soluble fraction of rhLOX-1-ECD instead of hsLOX-1 as the assay calibrator and immunogen. We estimated the amount of rhLOX-1-ECD by ultraviolet spectrophotometry, using the …