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Numerous biological or chemical substances have posed a serious threat to human beings and the demand for decentralized, point-of-care (POC) diagnostic systems has accordingly increased. Among many POC devices that are commercially available on the market, a personal glucose meter (PGM) with high portability, low cost, simple operation, and reliable quantitative ability has been regarded as one of the most successful. Recently, we developed a label-free and washing-free method for biomolecular detection using a personal glucose meter (PGM). ATP was selected as a model target, and cascade enzymatic reactions promoted by hexokinase and pyruvate kinase were adopted to link the amount of ATP to glucose that is detectable by a hand-held PGM. In principle, the presence of target ATP enables hexokinase to catalyze the conversion of glucose to glucose 6-phosphate, and thus the amount of glucose is decreased in proportion to the amount of ATP. The regenerated ATP from ADP is again supplemented to catalyze multiple rounds of cascade enzymatic reactions, leading to signal amplification. As a result, the change of glucose amount that is dependent to ATP amount is simply measured by a hand-held PGM. By employing this strategy, we successfully detected various targets containing nucleic acids, proteins, enzymes activity, small molecules with high sensitivity and selectivity even in real samples. Importantly, the developed system does not require expensive modification and washing steps but is conveniently operated with a commercially available PGM, which would pave the way for the development of a simple and cost-effective sensing platform.