Electrophoretically mediated microanalysis of beta-galactosidase on microchips.
Electrophoretically mediated microanalysis (EMMA) is a method of accomplishing chemical analyses, typically in an open-tubular capillary, due to the difference in the electrophoretic mobility between the particular reagents. This work reports on combining this technique onto microfabricated systems. Two methods of this technique were applied, constant potential and zero potential EMMA onto chips. A dosage response curve was run using this constant potential mode that resulted in a linear response over three orders of substrate concentration magnitude. The chemical system used here is beta-galactosidase (beta-Gal) as the enzyme and fluorescein mono-beta-D-galactopyranoside (FMG) as the substrate. The zero potential mode was used to amplify product turnover using various incubation times. Using this technique and a 10 min incubation, approximately 40000 enzyme molecules could be detected. The zero potential mode is also used in conjunction with an internal standard to show how one can quantitate using this method. The power and ease of utility of this technique is described.
Burke
B J
BJ
Department of Chemistry, Purdue University, West Lafayette, IN 47907, USA.
Regnier
F E
FE
D019075:Electrophoresis, Capillary / Q000379:methods; D007202:Indicators and Reagents; D007700:Kinetics; D008832:Microchemistry / Q000379:methods; D012680:Sensitivity and Specificity; D012984:Software; D001616:beta-Galactosidase / Q000032:analysis*
D007202:Indicators and Reagents; D001616:beta-Galactosidase