A micro-method for the rapid ultracentrifugal separation of human plasma high density lipoprotein subfractions, HDL2 and HDL3.

Currently available ultracentrifugal procedures for separating the two major density subfractions of HDL (HDL2 and HDL3) are time-consuming and expensive, and require relatively large volumes of plasma. In order to overcome these problems, a rapid and simple procedure, requiring only 0.5 ml plasma, has been developed. After precipitation of apoprotein B-containing lipoproteins with heparin and MnCl2, 120 microliters of the supernatant is adjusted to a density of 1.125 g/ml with aqueous NaCl-NaBr, and centrifuged for 3.5 h at 160000 x g in an air-driven bench-top ultracentrifuge (Beckman Airfuge). The HDL2 is then removed in a tube-slicer, constructed to permit six Airfuge tubes to be sliced simultaneously, and the cholesterol content assayed by a cholesterol oxidase procedure. The method provides an alternative to the recently described heparin/MnCl2/dextran sulphate double-precipitation procedure, with the advantage of permitting quantitative recovery of intact HDL2.