ELECTROPHORETIC SEPARATIONS OF PROTEINS IN CAPILLARIES WITH HYDROLYTICALLY STABLE SURFACE-STRUCTURES

A procedura for obtaining highly stable coated capillaries for use In caplłiary electrophoresis (CE) is described. Reaction of surface-chlorinated fused silica capiares with the Grignard reagent, vinyl magnesium bromide, followed by reaction of the vinyl group with acrylamide, results In an Immobilized layer of Polyacrylamide attached through hydrolytically stable Sl-C bonds. This method Is an extension of the capiliary coating procedure described previously by Hjerten, differing in the means by which the Polyacrylamide layer Is bonded to the capiliary walls. Capiliaries treated In the manner described here can be used over a pH range of 2–10.5, without noticeable decomposition of the coating. in comparison to uncoated capillaries, separations of proteins using such coated capillaries are improved due to a reduction in protein adsorption to the capillary walls, although interaction Is still present to some degree as evidenced by an inability to obtain plate counts as high as those predicted by theory. Electroosmotic flow is virtually eliminated in the coated capiliaris, resulting In improved reproductibilities of protein migration times in comparison to uncoated capillaries. Additionally, peak skew is evaluated for model proteins and improvements are noted for the coated capillaries. Results are presented for separations of model protein mixtures, comparing the performance of the vinyl-bound Polyacrylamide coated capillaries and uncoated capillaries at both high and low pH extremes. © 1990, American Chemical Society. All rights reserved.