Interface of an array of five capillaries with an array of one-nanoliter wells for high-resolution electrophoretic analysis as an approach to high-throughput chemical cytometry.
We report a system that allows the simultaneous aspiration of one or more cells into each of five capillaries for electrophoresis analysis. A glass wafer was etched to create an array of 1-nL wells. The glass was treated with poly(2-hydroxyethyl methacrylate) to control cell adherence. A suspension of formalin-fixed cells was placed on the surface, and cells were allowed to settle. The concentration of cells and the settling time were chosen so that there was, on average, one cell per well. Next, an array of five capillaries was placed so that the tip of each capillary was in contact with a single well. A pulse of vacuum was applied to the distal end of the capillaries to aspirate the content of each well into a capillary. Next, the tips of the capillaries were placed in running buffer and potential was applied. The cells lysed upon contact with the running buffer, and fluorescent components were detected at the distal end of the capillaries by laser-induced fluorescence. The electrophoretic separation efficiency was outstanding, generating over 750,000 theoretical plates (1,800,000 plates/m). In this example, AtT-20 cells were used that had been treated with TMR-G(M1). The cells were allowed to metabolize this substrate into a series of products before the cells were fixed. The number of cells found in each well was estimated visually under the microscope and was described by a Poisson distribution with mean of 0.98 cell/well. This system provides an approach to high-throughput chemical cytometry.
Boardman
Anna K
AK
Department of Chemistry, University of Washington, Seattle Washington 98195, USA.
McQuaide
Sarah C
SC
Zhu
Cuiru
C
Whitmore
Colin D
CD
Lidstrom
Mary E
ME
Dovichi
Norman J
NJ
D000818:Animals; D002460:Cell Line; D002518:Ceramides / Q000032:analysis / Q000378:metabolism; D019075:Electrophoresis, Capillary / Q000295:instrumentation / Q000379:methods*; D005434:Flow Cytometry / Q000295:instrumentation / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D005677:G(M1) Ganglioside / Q000032:analysis / Q000378:metabolism; D005678:G(M2) Ganglioside / Q000032:analysis / Q000378:metabolism; D005732:Gangliosides / Q000032:analysis / Q000378:metabolism*; D036103:Nanotechnology / Q000295:instrumentation / Q000379:methods; D012235:Rhodamines / Q000737:chemistry / Q000378:metabolism
D002518:Ceramides; D005456:Fluorescent Dyes; D005732:Gangliosides; D012235:Rhodamines; D005678:G(M2) Ganglioside; D005677:G(M1) Ganglioside; C005358:tetramethylrhodamine