Capillary electrophoresis of histone H1 variants at neutral pH in dynamically modified fused-silica tubing

Existing methods for the analysis of histone H1 by capillary electrophoresi s (CE) employ acidic buffers (pH <3.0) to suppress silanol ionization and m inimize the loss of these extremely basic proteins by adsorption to capilla ry walls. Here we describe the use of Polybrene (PB) as a dynamic modificat ion reagent in a simple procedure that facilitates the analysis of chicken H1 at neutral pH. PB is adsorbed to the inner surfaces of capillaries to re nder them cationic prior to use and a low concentration of PB is included i n the electrolyte to replenish the coating during use. Inclusion of ethylen ediaminetetraacetic acid (EDTA) in the electrolyte results in the assembly of a dynamic cation-exchange layer upon the immobilized PB that influences the relative mobilities of H1 variants. The six nonallelic variants of H1 k nown in this species as well as certain allelic variants are resolved. Beca use the procedure is effective in preventing the adsorption of proteins as basic as HI at neutral pH, this strategy should facilitate CE analyses of m any basic proteins under conditions that maintain their native conformation .