A Rapid Radioimmunoassay of Human Apolipoproteins C-Ii and C-Iii

Abstract

Apolipoprotein (apo) C-II is an activator of lipoprotein lipase, while apo C-III has the ability to inhibit apo C-II activated lipolysis. In order to study further the relationship between lipoprotein lipase mediated hydrolysis and the serum concentrations of apo C-II and apo C-III we have developed radioimmunoassays for these apolipoproteins.

Apo C-II and apo C-III were isolated from very low density lipoproteins (VLDL) by ion exchange chromatography. Apo C-II was further purified by affinity-chromatography. The purity of the two proteins was determined to be > 98% by polyacrylamide gel electrophoresis and immunodiffusion. The purified apolipoproteins were used as antigens, assay standards and labels.

Formalin-treated Staphylococcus aureus Cowan I was used for immunoprecipitation and were shown to give rapid uptake of immune complexes that could easily be harvested by centrifugation. The assays were shown to be sensitive (10 μg/l), specific, precise (inter- and intra-assay coefficients of variation below 10%), rapid (completed in less than 6 h) and simple to perform. Delipidation of serum and lipoproteins had no effect on the results, indicating that the immunologically active sites of apo C-II and apo C-III are exposed to the aqueous environment under assay conditions. Serum apo C-II and apo C-III levels of normolipidaemic subjects were approximately 25 mg/l and 110 mg/l respectively. Highly significant positive correlations were found between VLDL apo C-II and VLDL apo C-III, respectively, and VLDL triglycerides, VLDL cholesterol and total serum TG. There was also a highly significant correlation between the HDL cholesterol concentration and the HDL apo C-III concentration.

Key Words:

• apolipoproteins C-II and C-III
• lipoprotein
• radioimmunoassay