A conceptually new technique for fast DNA detection has been developed. Here, we report a fast and sensitive online fluorescence resonance energy transfer (FRET) detection technique for label-free target DNA. This method is based on changes in the FRET signal resulting from the sequence-specific hybridization between two fluorescently labelled nucleic acid probes and target DNA in a PDMS microfluidic channel. Confocal laser-induced microscopy has been used for the detection of fluorescence signal changes. In the present study, DNA hybridizations could be detected without PCR amplification because the sensitivity of confocal laser-induced fluorescence detection is very high. Two probe DNA oligomers (5′-CTGAT TAGAG AGAGAA-TAMRA-3′ and 5′-TET-ATGTC TGAGC TGCAGG-3′) and target DNA (3′-GACTA ATCTC TCTCT TACAG GCACT ACAGA CTCGA CGTCC-5′) were introduced into the channel by a microsyringe pump, and they were efficiently mixed by passing through the alligator teeth-shaped PDMS microfluidic channel. Here, the nucleic acid probes were terminally labelled with the fluorescent dyes, tetrafluororescein (TET) and tetramethyl-6-carboxyrhodamine (TAMRA), respectively. According to our confocal fluorescence measurements, the limit of detection of the target DNA is estimated to be 1.0 × 10-6 to 1.0 × 10-7 M. Our result demonstrates that this analytical technique is a promising diagnostic tool that can be applied to the real-time analysis of DNA targets in the solution phase.
Bibliographical noteFunding Information:
This work was supported by the Korea Science and Engineering Foundation (Grant numbers R01-2007-000-20238-0 and 2007-04431), the National Cancer Center of Korea (Grant number 0620400-1) and Seoul Research and Business Development Program (Grant number 10574).
- Confocal laser-scanning microscopy
- DNA hybridization
- Microfluidic sensor
- Real-time analysis
ASJC Scopus subject areas
- Biomedical Engineering