Measurement of high-density lipoprotein cholesterol in serum: comparison of six isolation methods combined with enzymic cholesterol analysis.

We compared six methods for isolating high-density lipoproteins from normal and lipemic serum. Cholesterol was determined by two different enzymic methods. The values for high-density lipoprotein cholesterol obtained with two (centrifugation-speed) modifications of the heparin/Mn2 method and with the phosphotungstate/Mg2t the dextran sulfate/Mg2, and the polyethylene glycol methods were compared with those obtained by ultracentrifugation. One cholesterol method, in which each determination was blank-corrected, was not interfered with by the various precipitation reagents. The other, in which there was no blank correction, was considerably interfered with by Mn2+ and dextran sulfate. Neither method was interfered with by phosphotungstate/Mg2 or by polyethylene glycol. Use of ethylenediaminetetraacetate in either cholesterol reagent was unhelpful (or deleterious). After correction for all interferences caused by the precipitation reagents and EDTA, values obtained for high-density lipoprotein cholesterol agreed well with those by ultracentrifugation (corrected for apo B-containing Iipoproteins in the d>1.063 kg/L fraction). Rocket immunoelectrophoresis showed no significant co-precipitation of high-density lipoproteins in any method. Various methods for complete removal of apo B-containing lipoproteins from lipemic serum appeared to vary widely in efficiency, but the optimized phosphotungstate method and the polyethylene glycol method appeared best. With them, only sera with triglyceride concentrations of 7.6 and 18.5 mmol/L or greater, respectively, could not be fractionated successfully. However, after ultrafiltration, accurate values were also obtained with the other methods. We conclude that the phosphotungstate method and the polyethylene glycol method are the best methods for HDL-cholesterol analysis when enzymic cholesterol analysis is used.