Isolation and Detection of Exosomal Mir210 Using Carbon Nanomaterial-Coated Magnetic Beads.

Abstract

MicroRNAs (miRNAs) are short non-coding RNAs that are found in various cellular compartments and play an important role in regulating gene expression. Extracellular miRNAs, such as those found within extracellular vesicles such as exosomes are involved in cell-to-cell communication. The intercellular transfer of miRNAs has been implicated in various diseases' pathogenesis including cancer and has been studied extensively as potential cancer biomarkers. However, the extraction of miRNA from exosomes is still a challenging task. The current nucleic acid extraction assays are expensive and labor-intensive. In this study, we demonstrated a microfluidic device for aptamer-based magnetic separation of the exosomes and subsequent detection of the miRNA using a fluorescence switching assay, which was enabled by carbon nanomaterials coated on magnetic beads. In the OFF state, the fluorophore-labelled cDNA is quenched using carbon nanomaterials. However, when the target miRNA210 is introduced, the cDNA detaches from the bead's surface, which leads to an increase in the fluorescence intensity (ON state). This increment was found to be proportional to miRNA concentration within the dynamic range of 0-100 nM with a detection limit of 5 pM. The assay was validated with spiked miRNA using the standard RT-PCR method. No notable cross-reactivity with other closely related miRNAs was observed. The developed method can be utilized for the minimally invasive detection of cancer biomarkers.