Friday, July 29, 2016

Dual-component gene detection for H7N9 virus – the combination of optical trapping and bead-based fluorescence assay

Di Cao, Cheng-Yu Li, Ya-Feng Kang, Yi Lin, Ran Cui, Dai-Wen Pang, Hong-Wu Tang

We present a strategy of dual-component gene detection for avian influenza A virus H7N9 by combining optical trapping and bead-based fluorescence bioassays. A low-cost 473 nm continuous DPSS laser, polystyrene (PS) beads with two different sizes (3 µm and 5 µm in diameter) and streptavidin-modified 605 nm quantum dots (SA-QDs) were exploited in this platform. The beads were employed to enrich the targets using the classic sandwich mode and tagged with the SA-QDs, then the QDs-tagged beads floating in the suspension were directly trapped and excited by the optical tweezers to give strong and stable fluorescence signal, which was applied to quantify the targets. The distinctive size information from the image of the trapped beads enabled the sorting of the different targets. The results show that tiny laser power 40 μW is applicable for both trapping and fluorescence excitation of the beads. Moreover, the limits of detection for hemagglutinin7 (H7) gene and neuraminidase 9 (N9) gene are 1.0 – 2.0 pM with good selectivity for the complex sample, which is two orders of magnitude lower than that of the conventional method. More importantly, this strategy was successfully used to identify the subtype of the avian influenza A virus by simultaneous detection of H7 and N9 gene sequences. The high sensitivity, good selectivity, typing ability and the low cost of the laser make this strategy a promising method for life science and clinical applications.

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