HYDROPHOBIC-INTERACTION CHROMATOGRAPHY OF PROTEINS ON CONTINUOUS BEDSDERIVATIZED WITH ISOPROPYL GROUPS

A new support is described for fast, high-resolution hydrophobic-inter action chromatography of proteins based on a continuous bed containing isopropyl groups. The bed is prepared by a simple procedure: a mixtur e of appropriate acrylamide monomers, including the monomer with the i sopropyl group, is polymerized directly in the chromatographic tube or , alternatively, in a beaker for columns with diameters greater than 1 -2 mm. The concentrations of the monomers were adjusted for optimum re solution and low flow resistance. Both standard (6 mm I.D.) and microb ore columns (0.015-0.320 mm I.D.) have been employed successfully for the separation of seven model proteins. A linear negative gradient of ammonium sulfate (2.2-0 M) in phosphate buffer was used for the elutio n. These columns share with all continuous bed columns the properties that the resolution is roughly independent of the flow-rate, that the gel is sufficiently rigid to give high flow-rates at moderate pressure s and that the efficiency for the separation of proteins is high and i ndependent of the bed height for beds longer than 4-5 cm. The residenc e time of the proteins on the column has no observable influence on th e appearance of the chromatograms.