A Highly Sensitive Strategy for Fluorescence Imaging of MicroRNA in Living Cells and in Vivo Based on Graphene Oxide-Enhanced Signal Molecules Quenching of Molecular Beacon

In situ imaging of microRNA (miRNA) in living cells and in vivo is beneficial for promoting the studies on miRNA-related physiological and pathological processes. However, the current strategies usually have a low signal to -background ratio, which greatly affects the sensitivity and imaging performance. To solve this problem, we developed a highly sensitive strategy for fluorescence imaging of miRNA in living cells and in vivo based on graphene oxide (GO) enhanced signal molecule quenching of a molecular beacon (MB). 2CyS-MB was designed by coupling two Cy5 molecules onto the opposite ends of MB. The fluorescence intensities of two Cy5 molecules were reduced because of the self quenching effect. After adsorbing on the GO surface, the fluorescence quenching of the molecules was enhanced by fluorescence resonance energy transfer. This double quenching effect significantly reduced the fluorescence background. In the presence of one miRNA molecule, the fluorescence signals of two Cy5 molecules were simultaneously recovered. Therefore, a significantly enhanced signal-to-background ratio was obtained, which greatly improved the detection sensitivity. In the presence of miRNA, the fluorescence intensity of 2Cy5-MB-GO recovered about 156 times and the detection limit was 30 pM. Compared with 1Cy5-MB-GO, the elevated fluorescence intensity was enhanced 8 times and the detection limit was reduced by an order of magnitude. Furthermore, fluorescence imaging experiments demonstrated that 2Cy5-MB-GO could visually detect microRNA-21 in various cancer cells and tumor tissues. This simple and effective strategy provides a new sensing platform for highly sensitive detection and simultaneous imaging analysis of multiple low-level biomarkers in living cells and in vivo.