Absolute Quantification of MicroRNAs in a Single Cell with Chemiluminescence Detection Based on Rolling Circle Amplification on a Microchip Platform

The absolute quantification of miRNAs in a single cell allows to better understand the heterogeneity of cells and the relationship between miRNAs and diseases. However, seldom methods for miRNA quantification in a single cell have been reported because the miRNA content in a single cell is very low. Herein, an ultrasensitive chemiluminescence assay strategy based on rolling circle amplification (RCA) on a microchip platform was proposed for the absolute quantification of miRNAs in a single cell. In this strategy, a ring probe with specificity was designed and synthesized, which could perform RCA for target miRNAs to improve the sensitivity and satisfy the need of absolute quantification of miRNAs in a single cell. The 20 liver cancer cells (HepG2) and 20 normal liver cells (HL-7702) were analyzed using this method; it is found that the miRNA-21 contents varied among cells, and miRNA-21 was overexpressed in HepG2 cells. Compared with traditional methods, the proposed strategy has many advantages such as low cost, simple operation, short analysis time, good specificity, and lower probability of false positives. This method is expected to be one of the powerful tools for the absolute quantification of miRNAs in a single cell.

Automated summary

This study proposed an ultrasensitive chemiluminescence assay strategy for the absolute quantification of miRNAs in a single cell, based on rolling circle amplification on a microchip platform. The method was able to detect differences in miRNA content between cells and identify overexpression of miRNA-21 in HepG2 cells. The strategy offers advantages such as low cost, simplicity, short analysis time, good specificity, and lower false positive rates compared to traditional methods.