Electrokinetic analyte transport assay for alpha-fetoprotein immunoassay integrates mixing, reaction and separation on-chip.
A rapid and highly sensitive CE immunoassay method integrating mixing, reaction, separation, and detection on-chip is described for the measurement of alpha-fetoprotein (AFP), a liver cancer marker in blood. Antibody-binding reagents, consisting of 245-bp DNA coupled anti-AFP WA1 antibody (DNA-WA1) and HiLyte dye-labeled anti-AFP WA2 antibody (HiLyte-WA2), and AFP-containing sample were filled into adjacent zones of a chip channel defined by the laminar flow lines of the microfluidic device using pressure-driven flow. The channel geometry was thus used to quantitatively aliquot the reagents and sample into the chip. DNA-WA1 was electrokinetically concentrated in the channel and sequentially transported through the AFP-sample zone and HiLyte-WA2 zone by ITP in such a manner that the AFP sandwich immune complex formation took place in the sample and HiLyte-WA2 zones. The sandwich AFP immune complex was then detected by LIF after CGE in a separation channel that was arranged downstream of the reaction channel. AFP was detected within 136 s with a detection sensitivity of 5 pM. The on-chip immunoassay described here, applying ITP concentration, in-channel reaction, and CGE separation, has the potential of providing a rapid and sensitive method for both clinical and research applications.
Kawabata
Tomohisa
T
New Diagnostics Business and Technology Development Department, Wako Pure Chemical Industries, Mountain View R&D Center, Mountain View, CA, USA.
Wada
Henry G
HG
Watanabe
Mitsuo
M
Satomura
Shinji
S
D046209:Electrophoresis, Microchip / Q000379:methods*; D007118:Immunoassay / Q000379:methods*; D012680:Sensitivity and Specificity; D000509:alpha-Fetoproteins / Q000032:analysis*
D000509:alpha-Fetoproteins