Simultaneous detection of multiple exosomal microRNAs for exosome screening based on rolling circle amplification.

ABSTRACT

Exosomal microRNAs (miRNAs) have attracted great attention as predictive and prognostic biomarkers of cancer. Profiling of miRNAs plays a key role in the effective diagnosis of cancers. However, simultaneous quantification of multiple miRNAs is challenging due to their homology and low abundance especially in exosomes. Here, we developed a sensitive detection method for multiple exosomal miRNAs with the help of rolling circle amplification (RCA). In contrast of the traditional ways, this method takes the advantages of both the multiplex sensing ability and the simplicity of RCA. Specifically, multiple exosomal miRNAs from different cell lines were replicated simultaneously through RCA and detected using designed molecular beacons (MBs). miRNA-21, miRNA-122 and miRNA-155 were chosen as the targets, which are overexpressed in cancers. Normalized fluorescence intensities of MB were used to imply the relative concentrations of these miRNAs. The obtained relative miRNAs expression levels could be used to distinguish the breast cancer exosome from normal one. If the varieties of the detected exosomal miRNAs are abundant enough, the concentration ratios of miRNAs could basically indicate the corresponding exosome and exosome screening could be realized. Such exosomal miRNA profiling and exosome screening can assist cancer diagnosis, which is promising in clinical application.