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This study introduces a new molecular beacon method for precisely detecting cell-cell fusion. This approach allows for sensitive, quantitative visualization and analysis of fusion events, aiding disease and infection research.

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Area of Science:

  • Biotechnology
  • Cell Biology
  • Molecular Diagnostics

Background:

  • Cell-cell fusion is crucial for understanding disease progression and viral infections.
  • Sensitive and quantitative detection of cell-cell fusion remains a significant experimental challenge.
  • Existing methods lack the precision required for detailed fusion analysis.

Purpose of the Study:

  • To develop a facile and precise molecular beacon (MB)-based method for quantitative detection of cell-cell fusion.
  • To establish a reliable system for visualizing and quantifying fusion events in real-time.
  • To enable the study of fusion dynamics in cells with varying receptor expression levels.

Main Methods:

  • Construction of virus-mimicking fusogenic effector cells (293-S-EGFP) by transfecting HEK 293 cells with spike protein (S protein) and enhanced green fluorescent protein (EGFP).
  • Silencing glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression in effector cells and delivering MBs for GAPDH mRNA detection.
  • Utilizing MBs that migrate and hybridize with target cell GAPDH mRNA upon fusion, inducing fluorescence emission.
  • Quantification of cell-cell fusion using fluorescence microscopy and flow cytometry.

Main Results:

  • The MB-based method successfully visualized and quantified cell-cell fusion events.
  • Fluorescence intensity directly correlated with the number of fused target cells.
  • The method differentiated fusion efficiencies in target cells expressing varying levels of angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2).

Conclusions:

  • The developed MB-based approach offers a convenient and efficient tool for the precise quantitative detection of cell-cell fusion.
  • This method facilitates the study of fusion mechanisms and can be applied to diverse cell types and experimental conditions.
  • The findings provide a valuable platform for advancing research in virology, immunology, and developmental biology.