A QUANTITATIVE IMMUNOASSAY FOR THE LYSINE-BINDING FUNCTION OF LIPOPROTEIN(A) - APPLICATION TO RECOMBINANT APO(A) AND LIPOPROTEIN(A) IN PLASMA

Apo(a), the unique apoprotein of lipoprotein(a) (Lp[a]), can express l ysine-binding site(s) (LBS). However, the LBS activity of Lp(a) is var iable, and this heterogeneity may influence its pathogenetic propertie s. An LBS-Lp(a) immunoassay has been developed to quantitatively asses s the LBS function of Lp(a). Lp(a) within a sample is captured with an immobilized monoclonal antibody specific for apo(a), and the captured Lp(a) is reacted with an antibody specific for functional LBS. The bi nding of this LBS-specific antibody is then quantified by using an alk aline phosphatase-conjugated disclosing antibody. The critical LBS-spe cific antibody was raised to kringle 4 of plasminogen. When applied to plasma samples, the LBS activity of Lp(a) ranged from 0% to 100% of a n isolated reference Lp(a); the signal corresponded to the percent ret ention of Lp(a) on a lysine-Sepharose column but did not correlate wel l with total Lp(a) levels in plasma. Mutation of residues in the putat ive LBS in the carboxy-terminal kringle 4 repeat (K4-37) in an eight-k ringle apo(a) construct resulted in marked but not complete loss of ac tivity in the LBS-Lp(a) immunoassay. These data suggest that this krin gle is the major but not the sole source of LBS activity in apo(a). Th e LBS-Lp(a) immunoassay should prove to be a useful tool in establishi ng the role of the LBS in the pathogenicity of Lp(a).