Capillary Gel Electrophoresis Separation of dsDNA Molecules Using Tetrahydroxyborate Stabilized Hydroxyethyl Cellulose – Polyvinylpyrrolidone Physical Copolymer

Abstract

Abstract The advent of nucleic-acid–based therapeutics has revived the need to develop efficient capillary gel electrophoresis methods for their rapid, high-resolution analysis. In this paper, we introduce a dual-network hydrogel-based physical copolymer, utilizing the excellent sieving properties of hydroxyethyl cellulose with interpenetrating polyvinylpyrrolidone, which also sufficiently suppresses electroosmotic flow. Boric acid in the background electrolyte served as a cross-linker/stabilizer for the hydroxyethyl cellulose-based separation matrix via the formation of spiroborane bridges. The addition of glycerol significantly increased the resolution of separations. The excellent sieving power of the tetrahydroxyborate-stabilized HEC-PVP physical copolymer is demonstrated by the separation of dsDNA restriction fragments in a wide base pair range of 67 to 10 000, even capable of resolving identical chain length fragments. As a practical example, plasmid DNA integrity monitoring was demonstrated, facilitating the path toward a novel high-resolution separation matrix for gene therapy products in the biopharmaceuticals industry.