Communications Biology (Apr 2023)

Single cell quantification of microRNA from small numbers of non-invasively sampled primary human cells

  • Vanessa Ho,
  • Jonathan R. Baker,
  • Keith R. Willison,
  • Peter J. Barnes,
  • Louise E. Donnelly,
  • David R. Klug

DOI
https://doi.org/10.1038/s42003-023-04845-8
Journal volume & issue
Vol. 6, no. 1
pp. 1 – 11

Abstract

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Abstract Expression levels of microRNAs (miRNAs) in single cells are low and conventional miRNA detection methods require amplification that can be complex, time-consuming, costly and may bias results. Single cell microfluidic platforms have been developed; however, current approaches are unable to absolutely quantify single miRNA molecules expressed in single cells. Herein, we present an amplification-free sandwich hybridisation assay to detect single miRNA molecules in single cells using a microfluidic platform that optically traps and lyses individual cells. Absolute quantification of miR-21 and miR-34a molecules was achieved at a single cell level in human cell lines and validated using real-time qPCR. The sensitivity of the assay was demonstrated by quantifying single miRNA molecules in nasal epithelial cells and CD3+ T-cells, as well as nasal fluid collected non-invasively from healthy individuals. This platform requires ~50 cells or ~30 µL biofluid and can be extended for other miRNA targets therefore it could monitor miRNA levels in disease progression or clinical studies.