IR-mediated PCR in microdevices is an established technique for rapid amplification of nucleic acids. In this report, we have expanded the applicability of IR-PCR to quantitative determination of starting copy number by integrating fluorescence detection during the amplification process. Placing the microfluidic device between an IR longpass filter and a hot mirror reduced the background to a level that enabled fluorescence measurements to be made throughout the thermal cycling process. The average fluorescence intensity during the extension step showed the expected trend of an exponential increase followed by a plateau phase in successive cycles. PUC19 template at different starting copy numbers were amplified and the threshold cycle showed an increase for decreasing amounts of starting DNA. Amplification efficiency was 80% and gel separation indicated no detectable non-specific product. A melting curve was generated using IR heating and indicated a melting temperature of 85 °C for the 304 bp amplicon which compared well to the melting temperature obtained using a conventional PCR system. This methodology will be applicable in other types of IR-mediated amplification systems, such as isothermal amplification and in highly integrated systems that combine pre- and post-PCR processes.

In practice, the application of the majority of carbon dots (CDs) has been limited in biomedical and bioimaging because of insufficient excitation/emission in near-infrared (NIR) regions. Near-infrared carbon dots (NIR-CDs) benefit from distinctive merits of low toxicity, strong biological penetration, minor susceptibility to the endogenous substances, and ameliorated NIR excitation/emission. Herein, multifunctional dual-emissive near-infrared carbon dots (dNIR-CDs) with intense NIR excitation/ emission are synthesized and subsequently utilized for the construction of a ratiometric nanosensor for rapid and sensitive detection of lysozyme (LYZ). When LYZ was exposed to the nanosensor, fluorescence quenching occurred quickly at the emission peak of 680 nm by the dynamic quenching mechanism, while the fluorescence intensity of the emission peak of 460 nm remained unchanged. The developed nanosensor exhibited a wide linear response within the range of 0.03−10 μM and a reproducible detection limit at the level down to 7 nM for LYZ. By virtue of the high selectivity and sensitivity of the nanosensor, LYZ can be detected at trace levels in complicated human urine samples. Furthermore, the high throughput of the nanosensor for cell imaging is demonstrated.

Micro total analysis systems (-TAS) or labs-on-achip, have been spreading rapidly due to their desirable characteristics, including reductions in reagent consumption, space requirements and analysis time. This work aimed at establishing an integrated microfluidic system which can supply the cells with fresh medium of oxygen and nutrition continuously at a control flow rate mimicking the microenvironment in vivo. Human non-small cell lung cancer cell line SPCA1 was seeded in a microchip supplied with fresh medium at a constant rate of 15 mm/24 h controlled by a pump. The expression of P-gp for verapamil-pretreated or non-pretreated cells was assayed with immunofluorescence. Both groups cells were exposed to anticancer drug VP-16 at 30 microM for 6 h before the apoptosis analysis online. The results indicated that the cells could grow and spread well for 4 days in the microfluidic system successively furnished with fresh medium. Immunofluorescence assay showed that the intensity of the fluorescence for the verapamil-pretreated cells was obvious weak compared with that of nonpretreated cells. Apoptosis analysis demonstrated that the percentage of apoptotic cells for verapamil-pretreated group increased around twofold compared with that of nonverapamil pretreated group (26.5+/-2.5% versus 10.9+/- 0.85%, p<0.05), showing a similar results as by flow cytometry analysis. All these indicate that P-gp plays an important role in the resistance to VP-16 in SPCA1, the microfluidic system provides a suitable environment for cells survival and is valuable in long time cell culture and bioassays mimicking the microenvironment in vivo and deserved to be studied further.

Novel fluorescent microfluidic paper chips were developed by the combination of molecularly imprinted polymers and microfluidic paper chips with CdTe quantum dots for the specific recognition and sensitive detection of the pesticide 2,4-D.