Light-Up Sensing Citrate Using a Capture-Selected DNA Aptamer

Citrate is a key metabolite and nutrient in humans. Its level is associated with many diseases from tumor growth to bone diseases. Detection of citrate has relied on its high negative charge, metal chelating properties and as an enzyme substrate. In this work, the capture-selection method is used to isolate DNA aptamers for citrate. After 18 rounds of selection, a highly converged library is obtained and the first two sequences reached 99.6% of the library. Using the most abundant sequence named CA1, thioflavin T fluorescence spectroscopy and isothermal titration calorimetry show dissociation constants of 7.4 and 4.4 µm citrate, respectively. CA1 does not require sodium for binding but requires 1.0 mm magnesium. Among the tested carboxylate molecules, only citrate can bind to the aptamer. A light-up fluorescence strand displacement biosensor is developed and it can detect citrate in simulated urine with a detection limit of 1.1 µm. This short 42-nucleotide aptamer can be readily adapted to other types of sensing mechanisms for the detection of citrate.