title: Simplified transient isotachophoresis/capillary gel electrophoresis method for highly sensitive analysis of polymerase chain reaction samples on a microchip with laser-induced fluorescence detection.
abstract: We present a sensitive, simple and robust on-chip transient isotachophoresis/capillary gel electrophoresis (tITP/CGE) method for the analysis of polymerase chain reaction (PCR) samples. Using chloride ions in the PCR buffer and N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) in the background electrolyte, respectively, as the leading and terminating electrolytes, the tITP preconcentration was coupled with CGE separation with double-T shaped channel network. The tITP/CGE separation was carried out with a single running buffer. The separation process involved only two steps that were performed continuously with the sequential switching of four voltage outputs. The tITP/CGE method showed an analysis time and a separation efficiency comparable to those of standard CGE, while the signal intensity was enhanced by factors of over 20. The limit of detection of the chip-based tITP/CGE method was estimated to be 1.1 ng/mL of DNA in 1x PCR buffer using confocal fluorescence detection following 473 nm laser excitation.
authors.affiliation: Bioengineering Center, Improve Medical Instrument's Co., Ltd., 510370 Guangzhou, China. ruark@126.com
authors.forename: Dayu
authors.initials: D
authors.lastname: Liu
authors.forename: Ziyou
authors.initials: Z
authors.lastname: Ou
authors.forename: Mingfei
authors.initials: M
authors.lastname: Xu
authors.forename: Lihui
authors.initials: L
authors.lastname: Wang
mesh_terms: D004247:DNA / Q000737:chemistry*; D004586:Electrophoresis / Q000379:methods*; D019075:Electrophoresis, Capillary / Q000379:methods*; D046209:Electrophoresis, Microchip / Q000379:methods*; D005453:Fluorescence; D007834:Lasers; D016133:Polymerase Chain Reaction; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity
chemical_list: D004247:DNA