Capillary electrophoresis chips with a sheath-flow supported electrochemical detection system.
Microfabricated capillary electrophoresis chips containing an integrated sheath-flow electrochemical detector are developed with the goal of minimizing the influence of separation voltages on end-column detection while maintaining optimum performance. The microdevice consists of an upper glass wafer carrying the etched separation, injection, and sheath-flow channels and a lower glass wafer on which gold- and silver-plated electrodes have been fabricated. The sheath-flow channels join the end of the separation channel from each side, and gravity-driven flow carries the analytes to the electrochemical detector placed at working distances of 100, 150, 200, and 250 microm from the separation channel exit. The performance of this detector is evaluated using catechol and a detection limit of 4.1 microM obtained at a working distance of 250 microm. Detection of DNA restriction fragments and PCR product sizing is demonstrated using the electroactive intercalating dye, iron phenanthroline. Additionally, an allele-specific, PCR-based single-nucleotide polymorphism typing assay for the C282Y substitution diagnostic for hereditary hemochromatosis is developed and evaluated using ferrocene-labeled primers. This study advances the feasibility of high-speed, high-throughput chemical and genetic analysis using microchip electrochemical detection.
Ertl
Peter
P
Department of Chemistry and Biophysics Graduate Group, University of California, Berkeley, CA 94720, USA.
Emrich
Charles A
CA
Singhal
Pankaj
P
Mathies
Richard A
RA
D004563:Electrochemistry; D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods; D046210:Microfluidic Analytical Techniques; D016133:Polymerase Chain Reaction / Q000379:methods; D012680:Sensitivity and Specificity; D013499:Surface Properties; D013997:Time Factors