Reversed-phase high-performance liquid chromatography of red blood cell membranes

Three reversed-phase (Hi-Pore RP-318, Hi-Pore RP-304, and Bio-Gel TSK Phenyl-RP+) and one hydrophobic-interaction (Bio-Gel TSK Phenyl-5PW) columns were used in a Bio-Rad chromatography system to separate the membrane proteins of human erythrocytes. A linear gradient, starting with 0.05% trifluoroacetic acid and ending with 95% acetonitrile and 0.05% trifluoroacetic acid was used. The four columns demonstrated slightly different selectivities for the proteins in ghosts. These profiles were further altered when ghosts were solubilized with 0.1% sodium dodecyl sulfate. The columns with less hydrophobic packings and larger pore sizes appear to be best suited for reversed-phase analyses of erythrocyte membrane proteins. Detergent solubilization was unnecessary for good resolution of the protein components.