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摘要
This paper describes the development of a paper-based microfluidic device that detects long DNA amplicons on the basis of hybridization reactions with a covalently immobilized DNA probe and biotin-labeled signal DNA strands, and chemiluminescent (CL) reactions catalyzed by a horseradish peroxidase (HRP)streptavidin conjugate. The paper-based microfluidic CL biosensor is developed for detection of 198-bp DNA fragments obtained by PCR amplification of the Listeria monocyto genes hlyA gene, where the CL intensity is detected by a simple, inexpensive CCD. To our knowledge, the combined use of the CCD sensing and paper-based microfluidic CL detection is first proposed here. The paper-based device includes wax-screen printed channels, and further explores origami-type folds to create an easy-to-operate valve that makes it possible to successively wash the detection zone in-between hybridization steps. Compared with other paper-based wax-screen-printing methods, the proposed method is faster and needs lower heating temperature. In our DNA biosensor, CL signals generated using a HRP-Iuminol-H2O2 system are heightened with p-iodine phenol (PIP) and detected with a CCD system. Under optical conditions, a linear range of 1.94 x 10(-1) pmol/L to 1.94 x 10(4) pmol/L is achieved and the limit of detection is found to be 6.3 x 10(-2) pmol/L. And, this detection limit is 3-5 orders of magnitude lower than those obtained from other CCD-based microfluidic CL DNA assays. The results show that the proposed integration of paper-based microfluidics, CL method and CCD sensing offers great promises in providing a highly-sensitive, reliable and cost-effective solution for gene biosensing applications.
- 出版日期2015-3-31
- 单位华南师范大学
