Lipids of human erythrocytes have been separated by silicic acid column chromatography, one-dimensional chromatography on silicic acid impregnated paper, or thin-layer chromatography (TLC), and combinations of silicic acid column chromatography with one-dimensional paper chromatography or TLC. The two-dimensional TLC procedure originally applied to brain lipid analysis (1) has been applied to erythrocyte lipids of various mammalian species (Z-4). In this report on human erythrocytes and in an accompanying paper on human plasma (5)) we describe improved resolution of human blood lipids by the use of two-dimensional TLC alone or combined with ion-exchange cellulose column chromatography.
METHODS
Separation and Washing of Erythrocytes
Blood was obtained from healthy human subjects after an overnight fast. Disodium ethylenediaminetetraacetate (EDTA, 1 mg/ml of whole blood) was used as anticoagulant. The 'blood was centrifuged for 15 min at 80g (4"(Z), the platelet-rich plasma (PRP) transferred to another tube, and the cells and PRP centrifuged at 15009 for 10 min (4ยฐC). The buffy coat and uppermost portion of the red cell mass were removed and the red cells washed three times with equal volumes of an isosmotic 423'