Real-Time PCR Using a PCR Microchip with Integrated Thermal System and Polymer Waveguides for the Detection of Campylobacter Jejuni

Abstract

A novel real-time PCR microchip platform with integrated thermal system and polymer waveguides has been developed. By using the integrated optical system of the real-time PCR chip, cadF -a virulence gene of Campylobacter jejuni, could specifically be detected. Two different DNA binding dyes, SYTOX Orange and TO-PRO-3, were added to the PCR mixture to realize the real-time PCR. The presented approach shows reliable real-time quantitative information of the PCR amplification of the targeted gene.

Micro real-time PCR device using a circulation pump

“…In contrast, the space-domain type utilizes a continuous flow while the three PCR steps maintain the predetermined temperatures (Kopp and Manz 1998;Yao et al 2005;Wang et al 2005Wang et al , 2006aLee and Wang 2006). One example of the space-domain type amplifies the template DNA through serpentine fluid channels located on three heating blocks.…”

Fluorescence detection by miniaturized instrumentation based on non-cooled CCD minicamera and dedicated for lab-on-a-chip applications

“…From this point of view, two main requirements have to be fulfilled: (1) optical transparency in the visible light region, necessary for the introduction of the fluorescence-inducing light into the detection area and the collection of fluorescence light from the area of interest within the chip, and (2) low autofluorescence of LOC material ensuring a minimal optical background signal. The chips can be made entirely as a multilayer construction of light-transparent materials, such as glass, PDMS, SU-8, COC or other polymers [11][12][13][14][15][16][17][18] . On the other hand, only the top cover of the chip can be made of these materials, whereas the rest of the chip is made of non-transparent materials such as silicon or ceramic.…”

“…As a result, the photodetector must be very sensitive and low noise. Commonly applied detectors are cooled photomultiplier tubes (PMT) 12,13,18 , semiconductor photodiodes 14,16,17 , rather Peltier's module cooled charge coupled devices (CCD) as sensing matrix in video cameras 11 or lines in spectrophotometers 15 (Table 1). In spite of the well-known measurement methodology and many established technical solutions of epifluorescence microscope-based fluorescence readout, the main advantage of this configuration is its easy adaptation for laboratory-scale LOC fluorescence measurement.…”

Lab-on-a-chip fluorescence detection with image sensor and software-based image conditioning

“…From point of view of LOC material, two main requirements have to be fulfilled: (1) optical transparency necessary for introduction of the fluorescence excitation light and collection of the fluorescence light from an area of interest within the chip, and (2) low autofluorescence of LOC's material to minimize optical background signal. The chips can be whole made as multilayer construction of light-transparent materials -like glass, PDMS, SU-8, COC or other polymers, or only a top cover of the chip is made of these materials whereas body of the chip is made of silicon or ceramic [4][5][6][7][8][9][10][11]. It is also possible to make LOC from non-transparent materials like low-temperature co-fired ceramics (LTCC) but an integration of optical waveguides enabling introduction of the fluorescence inducing light into a detection chamber and covered with glass or PDMS is necessary [12].…”

“…In the detection channel, fluorescence light emitted by the fluorochrome is collected by the microscope objective with proper magnification, and guided to a photodetector by passing through filter(s) and dichroic mirror(s) to exclude the excitation light. Common detectors are cooled photomultiplier tubes (PMT) [5,6,11], semiconductor photodiodes [7,9,10], rather cooled charge coupled devices (CCD) as sensing matrix in video cameras [4] or lines in spectrophotometers [8].…”