Lab on a single microbead: an ultrasensitive detection strategy enabling microRNA analysis at the single-molecule level

Detection of a single nucleic acid molecule is of great significance for both fundamental biochemistry studies and clinical diagnostics. By using microRNA (miRNA) as a model target, herein, we have developed a single-microbead-based sensing (SMBS) platform, which simply enables the detection of miRNA at the single-molecule level. In this strategy, an isothermal exponential amplification reaction (EXPAR) is rationally designed towards specific miRNAs and all products of the EXPAR are integrated onto a single microbead for signal amplification and fluorescence enrichment. This pushes the detection of miRNAs down to 1 aM in a 5 mu L sample, corresponding to 3 copies of the miRNA molecule. This new strategy also affords high selectivity and it is capable of distinguishing among homologous miRNA family members even with a single-base difference. Due to its ultrahigh sensitivity and selectivity, the proposed SMBS platform has been successfully applied to the detection of miRNA extracted from a single cell.