Radioimmunoassay of apolipoprotein a-ii

A double antibody radioimmunoassay was developed for apolipoprotein A-II (apo A-II), one of the two major apoproteins of human high density lipoproteins (HDL). Apo A-II contains two identical polypeptide chains linked by a disulfide bond. A specific antiserum was raised in sheep. Tracer apo A-II was radiolabeled with 125I by the Bolton-Hunter technique. Reduction and carboxymethylation of apo A-II approximately doubled its immunoreactivity. Since normal sera were found to contain small amounts of monomeric apo A-II, this represented one potential source of error in the radioimmunoassay. Inclusion of 0.1 M sodium cholate in the assay system, however, led to identical immunoreactivity of dimeric apo A-II and the reduced and carboxymethylated protein. Radioimmunoassay using sheep anti-apo A-II detected 75% to 85% of the apo A-II contained in serum or HDL. Masked antigenic sites could be exposed by organic solvent extraction or, more simply, by dilution of serum samples in the buffer containing 0.1 M sodium cholate. Serum levels of apo A-II were measured in a population of consecutively and prospectively selected free-living subjects between 30 to 69 years of age. Levels were significantly (p less than 0.005) higher in females (n = 201; 42.0 +/- 10.3 mg/dl) than in males (n = 189; 39.0 +/- 8.4 mg/dl). Serum apo A-II levels correlated significantly with HDL-cholesterol levels but less strongly than apo A-I, the other major HDL apoprotein.