| 1. |
- Mahmoudian, Laili, et al.
(författare)
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Microchip electrophoresis for detection of circle-to-circle amplification products towards sensitive and rapid DNA analysis
- 2007
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Ingår i: Chemistry Letters. - 0366-7022 .- 1348-0715. ; 36:3, s. 396-397
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Tidskriftsartikel (refereegranskat)abstract
- A new method for fast and precise analysis of circle-to-circle amplification (C2CA) products by microchip electrophoresis has been developed. Stable C2CA products were produced by applying a new enzymatic step to C2CA. Detection was carried out within 55s with RSD of migration time of 3.6% (it = 6) enabling reproducibility and high speed. A real sample of bacterial pathogen (V. Cholerae) at single nucleotide level was detected successfully based on this method.
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| 2. |
- Mahmoudian, Laili, et al.
(författare)
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Microchip electrophoresis for specific gene detection of the pathogenic bacteria V. cholerae by circle-to-circle amplification
- 2008
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Ingår i: Analytical Sciences. - 0910-6340 .- 1348-2246. ; 24:3, s. 327-32
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Tidskriftsartikel (refereegranskat)abstract
- We have developed a new method for a fast and precise analysis of circle-to-circle amplification (C2CA) product for specific gene detection by microchip electrophoresis. In this method, we have added a new enzymatic step to the C2CA reaction, which could be carried out isothermally at 37 degrees C. Compared to the original single-stranded DNA, the double-stranded DNA that is produced by this enzymatic reaction is more reliable for analysis by microchip electrophoresis. C2CA product was detected within 55 s with high reproducibility by this method which was successfully applied to the detection of 10-ng genomic DNA of the pathogenic bacteria Vibrio. cholerae within 110 s. Purification was found to be an indispensable step for the analysis of the C2CA product of genomic DNA samples.
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| 3. |
- Mahmoudian, Laili, et al.
(författare)
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Rolling circle amplification and circle-to-circle amplification of a specific gene integrated with electrophoretic analysis on a single chip
- 2008
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Ingår i: Analytical Chemistry. - : American Chemical Society (ACS). - 0003-2700 .- 1520-6882. ; 80:7, s. 2483-2490
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Tidskriftsartikel (refereegranskat)abstract
- We have developed an integrated platform for rolling circle amplification (RCA) and circle-to-circle amplification (C2CA) of circular probe (padlock probe) and subsequent microchip electrophoretic detection of a specific gene on a poly(methyl methacrylate) microchip. RCA and C2CA were successfully carried out at a steady temperature of 37 degrees C in the sample well of the microchip, and their respective product was detected on the same channel of the microchip, which was prefilled with a polymer separation matrix and fluorescent dye. Using a species-specific padlock probe for bacterial pathogen V. cholerae, a 25-ng bacterial genomic DNA could be detected in less than 65 min (including RCA and microchip electrophoresis) by this platform. Stable dsDNA C2CA product of genomic DNA for V. cholerae can be detected with the introduced integrated platform. Furthermore, the usefulness of this technique for the monitoring of RCA was demonstrated. This integrated platform provides a sensitive, fast, high-throughput, and reproducible method for signal amplification and detection of the padlock probes in the same microchip and is a promising tool for highly specific gene detection strategies.
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| 4. |
- Onoshima, Daisuke, et al.
(författare)
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A deep microfluidic absorbance detection cell replicated from a thickly stacked SU-8 dry film resist mold
- 2012
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Ingår i: Analytical Methods. - : Royal Society of Chemistry (RSC). - 1759-9660 .- 1759-9679. ; 4, s. 4368-4372
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Tidskriftsartikel (refereegranskat)abstract
- Dry film resist SU-8 was used to make a thick mold for soft lithography of a poly(dimethylsiloxane) (PDMS) microfluidic chip with deep channels. The stacking of the SU-8 film enabled an ultra-thick (up to 500 μm) resist process on Si wafer. This process was fast and highly reproducible compared with the conventional liquid SU-8 process. The deep channel in the PDMS chip was utilized as a micro-flow cell for sensitive absorbance measurement. Sunset Yellow FCF dye was used to demonstrate absorption spectroscopy in the deep channel. Since the channel depth was proportional to the optical path length, which was proportional to the absorbance value, the PDMS chip achieved a detection limit (15.9 μM) comparable to U- or Z-shaped microfabricated absorbance detection cells in glass. Calibration curves for different solution concentrations were obtained with good R2 values (∼1). © 2012 The Royal Society of Chemistry.
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| 5. |
- Wang, Jun, 1976, et al.
(författare)
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Microfluidic biosensor for the detection of DNA by fluorescence enhancement and the following streptavidin detection by fluorescence quenching
- 2014
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Ingår i: Biosensors and Bioelectronics. - : Elsevier BV. - 0956-5663 .- 1873-4235. ; 51, s. 280-285
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Tidskriftsartikel (refereegranskat)abstract
- We reported an optical DNA/protein microfluidic sensor which consists of single stranded (ss) DNA-Cy3 probes on gold surface and simple line-shape microfluidic channel. These ssDNA-Cy3 probes with random sequence in bulk solution or on gold surface exhibits fluorescence enhancement after binding with complementary ssDNA (cssDNA) targets. Particularly it did not require complicated design or hairpin-like stem-loop conformation, which made it easier to be made and applied in analytes detection by fluorescence switching techniques. Using ssDNA-cy3 probes attached on gold surface in a microfluidic channel, strong fluorescence enhancement was measured by ssDNA with cssDNA binding or ssDNA with cssDNA-biotin binding. The following introduction of streptavidin resulted in fluorescence quenching (fluorescence decrease) because of the binding of hybridized DNA-biotin with streptavidin. This sensor showed strong affinity and high sensitivity toward the streptavidin, the minimum detectable concentration for streptavidin was 1 pM, equating to an absolute detection limit of 60 amol in this microfluidic channel. Microfluidic channel height and flow rate is optimized to increase surface reaction efficiency and fluorescence switching efficiency. In contrast to previously reported optical molecular beacon approach, this sensor can be used not only for the detection of cssDNA target, but also for the detection of streptavidin. This microfluidic sensor offers the promise of analyzing kinds of molecular targets or immunoreactions.
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| 6. |
- Yasui, Takao, et al.
(författare)
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Label-free detection of real-time DNA amplification using a nanofluidic diffraction grating
- 2016
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Ingår i: Scientific Reports. - : Springer Science and Business Media LLC. - 2045-2322. ; 6
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Tidskriftsartikel (refereegranskat)abstract
- Quantitative DNA amplification using fluorescence labeling has played an important role in the recent, rapid progress of basic medical and molecular biological research. Here we report a label-free detection of real-time DNA amplification using a nanofluidic diffraction grating. Our detection system observed intensity changes during DNA amplification of diffracted light derived from the passage of a laser beam through nanochannels embedded in a microchannel. Numerical simulations revealed that the diffracted light intensity change in the nanofluidic diffraction grating was attributed to the change of refractive index. We showed the first case reported to date for label-free detection of real-time DNA amplification, such as specific DNA sequences from tubercle bacilli (TB) and human papillomavirus (HPV). Since our developed system allows quantification of the initial concentration of amplified DNA molecules ranging from 1 fM to 1 pM, we expect that it will offer a new strategy for developing fundamental techniques of medical applications.
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