Study of Effect of Incubation Time and Temperature on the Linear Polyacrylamide-Coating Performance for the Separation of Proteins by Capillary Zone Electrophoresis

Abstract

The preparation of linear polyacrylamide (LPA)-coated capillary is a multistep, laborious process. In this study, five different LPA-coating procedures were compared regarding the capillary pretreatment (incubation time and temperature), the formation of the intermediate silyl layer and the attachment of the polymer top layer. LPA coatings were examined by the analysis of a standard mixture solution of insulin, rituximab and hemoglobin using a background electrolyte (BGE) of 50 mM HCOOH (pH 2.6). The results were contrasted with those that were obtained using the bare fused-silica capillary. The reproducibility of the simplest LPA-coating preparation was tested. The long-term stability of the capillary coating was evaluated through inter-day precision data using a large number of injections of the three-protein test mixture solution. The implementation of a short-term procedure at elevated temperature, guided by a comprehensive review of various conditions documented in the literature, yielded an efficiently functional LPA coating.