Multicolor fluorescence detection on an electrophoretic microdevice using an acoustooptic tunable filter.
An acoustooptic tunable filter (AOTF) is used to detect multiple fluorescent signals on a fluidic microdevice. A confocal laser-induced fluorescence detection setup is used to excite fluorescent dyes in glass microchannels, presenting a streamlined and robust detection system consisting of the narrow-bandwidth AO filter and a single photodetector. The flexibility of the filter is demonstrated by alternating between wavelengths for precise microchannel alignment and sweeping through a range of wavelengths for preliminary spectral characterization of subnanoliter probe volumes of target analytes. The AOTF is also coupled with an electrophoretic separation for the multicolor detection of PCR-amplified DNA against a labeled sizing standard, the discrimination of multiple amplicons overlapped in time, and the identification of amplified biowarfare agents in a fluorescent spiking experiment. Finally, to demonstrate the multicolor capability of the system, 19-wavelength detection is performed during the separation of a three-dye sample mixture.
Karlinsey
James M
JM
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904, USA.
Landers
James P
JP
D000162:Acoustics; D046209:Electrophoresis, Microchip / Q000295:instrumentation / Q000379:methods*; D005453:Fluorescence; D005456:Fluorescent Dyes / Q000032:analysis*; D012680:Sensitivity and Specificity; D013997:Time Factors
D005456:Fluorescent Dyes