Aptamer-linked in vitro expression assay for ultrasensitive detection of biomarkers

Abstract Signal amplification is a key step that determines the sensitivity of molecular assays. Although studies on aptamers have mostly focused on their target-binding ability, taking advantage of the gene-coding function of nucleic acids, we demonstrate here that aptamers can be engineered into diagnostic reagents that can both recognize a target and generate highly amplified detection signals. We developed a strategy that employs a ‘readable’ aptamer that consists of a single-stranded aptamer and a double-stranded reporter gene. After binding to its target via the aptamer region, the reporter gene of the readable aptamer produces amplified number of signal-generating enzymes through a subsequent in vitro expression reaction. In contrast to conventional enzyme-conjugation methods, this method allows the generation of far more amplified detection signals, thereby markedly increasing the sensitivity of detection enough to analyze a target present in aM concentrations.