Ultra small reaction chamber array for single-molecule studies of biological reaction

Summary form only given. We developed reaction chambers of micro-meter size to monitor biological reactions at single-molecule level. Micro-meter holes were fabricated on a silicone (poly-dimethylsiloxane, PDMS) sheet surface. Sample solutions were sandwiched between the fabricated sheet and a glass surface. Resultantly, solutions were enclosed in each chamber hermetically. The uniformly shaped chambers allow us stable and reversible enclosure of biological samples including ATP, protein, DNA strands and living cells. Due to its extremely small volume down to 1 femtoliter (a cube of 1 micron), very small number of molecules (several hundreds molecules) can reach up to detectable concentrations, micro-molars. When a commercially available enzyme assay mixtures were enclosed, detections of single enzyme molecules were readily achieved for beta-galctosidase and horse radish peroxidase without any elaborate optical systems. Reaction efficiencies of a molecular motor, F₁ATPase, were also measured at single-molecule level. These experiments certify large potential of the micro reaction chamber for high throughput and high sensitive analysis. Because this technology is based on a very simple logic and simple techniques, it would be compatible with mostly any other micro chip technologies. The other advantage of this technology is its cheapness; once the micro patterned silicon chip is prepared, micro patterned sheets can be reproduced using the chip as a mold. Thus, this novel and simple technique offers new highly sensitive and cheap detections.