Selection and characterization of dimethylindole red DNA aptamers for the development of light-up fluorescent probes

Abstract To develop novel label-free light-up probes with improved performance characteristics and low background, we selected DNA aptamers for dimethylindole red (DIR) by a modified affinity chromatography based SELEX method. DIR is an anionic propylsulfonate substituted red-emitting dye derivative of thiazole orange and exhibited weak fluorescence in fluid solution and in the presence of dsDNA. After 14 rounds of selection, a shortened 42-mer DNA aptamer with sub-micromolar dissociation constant (K d =0.65±0.17 μM) was selected. The fluorescent intensity of DIR was dramatically enhanced in the presence of the specific aptamer. The aptamer gave a 140-fold fluorescence enhancement in a saturated concentration. The DIR-aptamer pair could be potentially used as novel light-up fluorescent probe to construct sensors for various applications.