Parallel electrophoretic analysis of segmented samples on chip for high-throughput determination of enzyme activities.
Capillary electrophoresis (CE) on microfabricated structures has achieved impressive sample throughput by combining fast separation speed and parallel operations. One obstacle to further increasing throughput has been lack of methods for loading and injecting individual samples at a rate that matches analysis speed. To address this issue, we have developed a microfluidic device in which samples stored as nanoliter volume plugs segmented by a fluorocarbon oil are introduced sequentially to an array of three electrophoresis channels. A microfluidic interface consisting of patterned surface chemistry and geometric restriction was used to extract samples from each segmented flow channel and transfer to the respective electrophoresis channel for separation. Fluorescence detection was achieved by imaging the chip using a fluorescence microscope equipped with a charge-coupled device. Characterization of the system shows that injection volume is controlled by sample plug volume, flow rate during introduction, and voltage applied to the electrophoresis channel. The system was tested for a GTPase assay. Peak area ratios of enzyme product and internal standard had 6% relative standard deviations. Cross-contamination between peaks was 7%. Throughput of 120 samples in 10 min was achieved. Further development of the system may allow application to high-throughput applications such as drug screening.
Pei
Jian
J
Department of Chemistry, University of Michigan, Ann Arbor, Michigan 48109-1055, United States.
Nie
Jing
J
Kennedy
Robert T
RT
D055808:Drug Discovery; D019075:Electrophoresis, Capillary / Q000295:instrumentation*; D057075:Enzyme Assays / Q000295:instrumentation*; D020558:GTP Phosphohydrolases / Q000378:metabolism; D007267:Injections; D046210:Microfluidic Analytical Techniques; D013997:Time Factors
D020558:GTP Phosphohydrolases