Analysis of lipoproteins by microchip electrophoresis with high speed and high reproducibility

A method for the fast analysis of lipoproteins by microchip electrophoresis with light-emitting diode confocal fluorescence detection has been developed. Lipoproteins labeled with BODIPY FL C(5)-ceramide are found to strongly adsorb on the bare surface of a poly(methyl methacrylate) (PMMA) microchip. Sodium dodecyl sulfate and cetyltrimethylammonium bromide were therefore utilized to alter lipoproteins and channel surface to make them bear the same type of charge. After modification, the peak shape of lipoproteins was greatly improved, demonstrating lipoprotein adsorption on a PMMA chip dramatically reduced due to electrostatic repulsion. In addition, polymers were added into the running buffer to suppress electroosmotic flow and to serve as a sieving matrix. As a result, lipoprotein separation was manipulated by both electrophoretic mobilities and particle sizes. Various separation parameters including surfactant concentration, buffer pH, and polymer concentration as well as on-line concentration were investigated systematically. Under optimal conditions, two baseline separations of standard lipoproteins including high-density lipoprotein, low-density lipoprotein, and very low-density lipoprotein were achieved with different selectivity. This method affords high separation speed (within 100 s) and high reproducibility. The intraassay and interassay RSDs of lipoprotein migration times were in the range of 0.90-1.9%, indicating this method is highly reliable.