Theophylline is a potent bronchodilator with a narrow therapeutic index. A simple fluorescent biosensor that detects clinically relevant theophylline concentrations has been developed using the well-characterized theophylline binding RNA aptamer. Hybridization of the RNA aptamer to a fluorescently labeled DNA strand (FL-DNA) yields a fluorescent RNA:DNA hybrid that is sensitive to theophylline. The biosensor retains the remarkable selectivity of the RNA aptamer for theophylline over caffeine and is sensitive to 0-2 muM theophylline, well below the clinically relevant concentration (5-20 mg/L or approximately 10-50 muM). Adding a dabcyl quenching dye to the 3'-terminus of the fluorescently labeled DNA strand yielded a dual-labeled DNA strand (FL-DNA-Q) and increased the dynamic range of this simple biosensor from 1.5-fold to 4-fold.
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