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Instrumentation for Medium-Throughput Two-Dimensional Capillary Electrophoresis with Laser-Induced Fluorescence Detection

2006; American Chemical Society; Volume: 79; Issue: 2 Linguagem: Inglês

10.1021/ac061652u

1520-6882

Cuiru Zhu, Xinya He, James R. Kraly, Megan R. Jones, Colin D. Whitmore, David González‐Gómez, Michael J. Eggertson, Wes W. C. Quigley, Anna K. Boardman, Norman J. Dovic̀hi,

Innovative Microfluidic and Catalytic Techniques Innovation

In two-dimensional capillary electrophoresis, a sample undergoes separation in the first dimension capillary by sieving electrophoresis. Fractions are periodically transferred across an interface into a second dimension capillary, where components are further resolved by micellar electrokinetic capillary electrophoresis. Previous instruments employed one pair of capillaries to analyze a single sample. We now report a multiplexed system that allows separation of five samples in parallel. Samples are injected into five first-dimension capillaries, fractions are transferred across an interface to 5 second-dimension capillaries, and analyte is detected by laser-induced fluorescence in a five-capillary sheath-flow cuvette. The instrument produces detection limits of 940 ± 350 yoctomoles for 3-(2-furoyl)quinoline-2-carboxaldehyde labeled trypsin inhibitor in one-dimensional separation; detection limits degrade by a factor of 3.8 for two-dimensional separations. Two-dimensional capillary electrophoresis expression fingerprints were obtained from homogenates prepared from a lung cancer (A549) cell line, on the basis of capillary sieving electrophoresis (CSE) and micellar electrophoresis capillary chromatography (MECC). An average of 131 spots is resolved with signal-to-noise greater than 10. A Gaussian surface was fit to a set of 20 spots in each electropherogram. The mean spot width, expressed as standard deviation of the Gaussian function, was 2.3 ± 0.7 transfers in the CSE dimension and 0.46 ± 0.25 s in the MECC dimension. The standard deviation in spot position was 1.8 ± 1.2 transfers in the CSE dimension and 0.88 ± 0.55 s in the MECC dimension. Spot capacity was 300.