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Photopolymerized acrylamide gel microarray used to immobilize the molecular beacon for label free DNA hybridization detection is described in this paper. Polyacrylamide gel microarray was prepared by UV photopolymerization of 4% acrylamide in 40% glycerol, 0.002%methylene blue, 0.012% TEMED and 0.1M phosphate buffer (PHequals7) with a Relpel-silane pretreated quartz mask. This kind of three-dimensional gel microarray provides more than 100 times great immobilization capacity. The hybridization and other processes with it resemble a homogeneous liquid phase reaction rather than a heterogeneous liquid-solid interface reaction. The specially designed molecular beacons contain a 15 base loop sequence with 5 base pair stem, a 20 base thymine as spacer, a 5'-end amino group for immobilization, a fluorescein in the middle of the sequence as the fluorophore and a 3'-end DABCYL as the quencher. Between the 5'-end amino group and the stem, the 20 base thymine is used to minimize destability caused by 5'-end immobilization. Confocal microscope was used to investigate the fluorescence intensity of gel immobilized molecular beacon probes. After hybridization we can easily distinguish complementary and noncomplementay targets with gel-immobilized molecular beacon probes. Image analysis showed that the fluorescence intensity ratio of complementary to noncomplementay probes is great than 5. The potential applications of gel-immobilized molecular beacon microarray are mutation detection, pathogenic detection, etc. in a parallel, cost saving and label-free detection way.
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