title: A simple and precise method for measuring HDL-cholesterol subfractions by a single precipitation followed by homogenous HDL-cholesterol assay.
abstract: HDL consists of two major subfractions, HDL2 and HDL3. This paper describes a simple method for assaying HDL subspecies by combining a single precipitation with a direct high density lipoprotein-cholesterol (HDL-C) assay. A precipitation reagent (0.06 ml) containing 1,071 U/ml heparin, 500 mmol/l MnCl2) and 12 mg/ml dextran sulfate was added to a serum (0.3 ml). The sample was incubated and centrifuged at 10,000 rpm for 10 min. HDL3-C was measured by a homogenous HDL-C assay in the supernatant, and HDL2-C was estimated by subtracting the HDL3-C from the direct HDL-C. The HDL3-C and HDL2-C values determined by the precipitation method were identical to those determined by ultracentrifugation, and there were excellent correlations between the methods in the measurements of HDL3-C and HDL2-C (r = 0.933 and 0.978, respectively; n = 102). The two methods also proved to be highly correlated in the measurement of apolipoprotein A-I and A-II in HDL subfractions. The HDL-C subfractions determined by ultracentrifugation were more closely associated with the homogenous HDL-C assay than with the total cholesterol assay, especially in the hypertriglyceridemic samples. Our method is far simpler and more precise than the classical dual precipitation method for HDL-C subfractions, and it can be easily performed in a routine chemical laboratory.
authors.affiliation: First Department of Internal Medicine, Showa University, School of Medicine, Tokyo, Japan. hirano@med.showa-u.ac.jp
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Hirano
authors.forename: Kyoko
authors.initials: K
authors.lastname: Nohtomi
authors.forename: Shinji
authors.initials: S
authors.lastname: Koba
authors.forename: Ayako
authors.initials: A
authors.lastname: Muroi
authors.forename: Yasuki
authors.initials: Y
authors.lastname: Ito
mesh_terms: D008076:Cholesterol, HDL / Q000097:blood / Q000145:classification* / Q000302:isolation & purification*; D006493:Heparin; D006801:Humans; D012016:Reference Values; D014461:Ultracentrifugation
chemical_list: D008076:Cholesterol, HDL; D006493:Heparin