Single Molecule Fluorescent Colocalization of Split Aptamers for Ultrasensitive Detection of Biomolecules

Single-molecule fluorescence imaging is a promising strategy for biomolecule detection. However, the accuracy of single-molecule method is often compromised by the false-positive events at the ultralow sample levels that are caused by the non-specific adsorption of the fluorescent labeled probe and other fluorescent impurities on the imaging surface. Here, we demonstrate an ultrasensitive single molecule detection assay based on dual-color fluorescent colocalization of spilt aptamers that was implemented to the measurement of adenosine triphosphate (ATP). The ATP aptamer was split into two fragments and labeled with green and red dye molecules respectively. When the two probes of split aptamers were brought together by the target ATP molecule, the two colors of fluorescence of two probes were simultaneously detected through two channels and projected to the correlated locations in the two halves of image. The colocalizaiton imaging of two split apatamer probes greatly excluded the false detection of biomo...