Red diode laser induced fluorescence detection with a confocal microscope on a microchip for capillary electrophoresis.
A highly sensitive laser induced fluorescence (LIF) detection system based on a 635 nm laser diode and cyanine-5 (Cy-5) dye, is described for use with a planar, microfluidic, capillary electrophoresis (CE) chip. The CE-chip is able to determine a protein biological threat agent simulant, ovalbumin (Ov), by performing an immunoassay separation of Cy-5 labeled anti-ovalbumin from its complex with Ov, in under 30 s. A confocal, epiluminescent detection system utilizing a photomultiplier tube gave optimum results with a 400 microm pinhole, an Omega 682DF22 emission filter, a 645DRLP02 dichroic mirror, a 634.54 +/- 5 nm excitation filter, and a Power Technology ACMO8 635 nm laser operated at 11.2 mW. Using this detector, a microchip CE device with a separation efficiency of 42,000 plates and an etch depth of 20 microm, gave a concentration detection limit of 9 pM Cy-5. This limit corresponds to the determination of 4560 injected molecules and detection of 900 of these molecules, given a probe volume of 1.6 pl and a probing efficiency of 20%.
Jiang
G
G
Department of Chemistry, University of Alberta, Edmonton, Canada.
Attiya
S
S
Ocvirk
G
G
Lee
W E
WE
Harrison
D J
DJ
D019075:Electrophoresis, Capillary / Q000295:instrumentation* / Q000379:methods; D005453:Fluorescence; D007834:Lasers; D018613:Microscopy, Confocal / Q000295:instrumentation / Q000379:methods