Precise Detection on Cell–Cell Fusion by a Facile Molecular Beacon-Based Method
| Autor: | Chang Xu, Xiao-He Ren, Di Han, Yan Peng, Jin-Ju Lei, Luo-Xiao Yu, Ling-Juan Liu, Wei-Chao Xu, Si-Xue Cheng |
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| Rok vydání: | 2022 |
| Předmět: | |
| Zdroj: | Analytical Chemistry. 94:17334-17340 |
| ISSN: | 1520-6882 0003-2700 |
| Popis: | Cell-cell fusion studies provide an experimental platform for evaluating disease progression and investigating cell infection. However, to realize sensitive and quantitative detection on cell-cell fusion is still a challenge. Herein, we report a facile molecular beacon (MB)-based method for precise detection on cell-cell fusion. By transfection of the spike protein (S protein) and enhanced green fluorescent protein (EGFP) in HEK 293 cells, the virus-mimicking fusogenic effector cells 293-S-EGFP cells were constructed to interact with target cells. Before mixing the effector cells with the target cells, the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression in 293-S-EGFP cells was silenced, and the MB for GAPDH mRNA detection was delivered into the GAPDH silenced 293-S-EGFP cells. Once cell-cell fusion occurred, MB migrated from the GAPDH silenced effector cells to the target cells and hybridized with GAPDH mRNA in the target cells to induce fluorescence emission. The cell-cell fusion can be easily visualized and quantitated by fluorescence microscopy and flow cytometry. The fluorescence intensity is strongly dependent on the number of fused target cells. This MB-based method can easily identify the differences in the cell fusions for various target cells with different angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) expression levels, resulting in dramatically different fluorescence intensities in fused target cells. Our study provides a convenient and efficient quantitative detection approach to study cell-cell fusion. |
| Databáze: | OpenAIRE |
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