Isolation and quantitation of apolipoproteins A-I and A-II from human high-density lipoproteins by fast-protein liquid chromatography.

Apolipoproteins (Apo) A-I and A-II from human high-density lipoproteins (HDL) were isolated and quantified by fast-protein liquid chromatography using a Superose 12 column (gel filtration) followed by a Mono Q column (anion-exchanger). The separation times were 45 min and 15 min, respectively. Identities of both apolipoproteins were confirmed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and by double immunodiffusion. Peak areas after Mono Q chromatography increased linearly with concentration for samples from 0.5 to 10 mg of protein with a mean ratio of Apo A-I to Apo A-II of 3.46 +/- 0.71 in normolipoproteinemic subjects. This precise technique is an alternative for preparing and quantifying both HDL apolipoproteins.