Cellular Physiology and Biochemistry (Nov 2018)

MicroRNA Quantitation During Dendritic Cell Endocytosis Using Imaging Flow Cytometry: Key Factors and Requirements

  • Qiang Tong,
  • Ying Zhu,
  • Dandan Zhang,
  • Qing Cai,
  • Wenchun Qu,
  • Christopher D. O. Cooper,
  • Jiaqi Wang,
  • Patrick C. McHugh

DOI
https://doi.org/10.1159/000495333
Journal volume & issue
Vol. 51, no. 2
pp. 793 – 811

Abstract

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Background/Aims: MicroRNA (miRNA)-induced suppression of dendritic cells (DCs) has been implicated in many diseases. Therefore, accurate monitoring of miRNA endocytosis by DCs is important for understanding the role of miRNAs in many diseases. Recently, a method for measuring the co-localization of Argonaute 2 (AGO2)-associated miRNAs on laser-scanning confocal microscopy method was proposed to localize the miRNAs. But its definition was limited by the number of observed cells through its accuracy. Methods: In this study, a method based on imaging flow cytometry was developed to localize miR-590-5p with fluorescent probes in DCs. miR-590-5p proven to play an important role in tumor immunity. This method enabled the quantification, visualization and localization of the fluorescence intensity in 30,000 individual cells. Results: Using this method, the DCs with different endocytotic ability were distinguished. The behaviour of miR-590-5p during endocytosis under the stimulation of tumor antigen in DCs was observed, binding to its cognate target mRNA and degradation in DCs. Conclusion: This method based on imaging flow cytometry provide an additional method to study miRNA processing in DCs, which makes it a valuable addition to existing miRNA research techniques.

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