Department of Cell Biology, University of Oklahoma Health Sciences Center, College of Medicine, 940 Stanton L. Young Blvd, Biomedical Sciences Building, Rm 553, Oklahoma City, OK, 73104, USA.
Cytotechnology. 2008 Nov;58(3):113-8. doi: 10.1007/s10616-009-9186-z. Epub 2009 Feb 28.
Cell-cell fusion is an important biological and pathological event. There are limited techniques for studying both the process of cell-cell fusion and the fate of fused cells. We have developed a non-invasive assay for the temporal analysis of cell-cell fusion, quantification of fused cells, and isolation of fused cells. Briefly, cells are transfected with either the T7 bacteriophage RNA polymerase, or yellow fluorescent protein (YFP) driven by a T7 specific promoter. Cells are mixed and induced to fuse. When cells expressing T7 RNA polymerase and T7 promoter driven YFP (T7-YFP) fuse and the cellular contents mix, the YFP is expressed. These YFP-positive cells can be detected with a fluorescent microscope, quantified by flow cytometry, or collected using fluorescence associated cell sorting. Isolated YFP-positive cells can be monitored to determine the fate of fused cells, specifically for the rates of growth, transformation, and changes in chromosome number.
细胞融合是一个重要的生物学和病理学事件。目前用于研究细胞融合过程和融合细胞命运的技术有限。我们开发了一种非侵入性的分析方法，用于对细胞融合进行时间分析、融合细胞的定量以及融合细胞的分离。简要地说，用 T7 噬菌体 RNA 聚合酶或黄色荧光蛋白（YFP）转染细胞，后者由 T7 特异性启动子驱动。将细胞混合并诱导融合。当表达 T7 RNA 聚合酶和 T7 启动子驱动的 YFP（T7-YFP）的细胞融合并且细胞内容物混合时，表达 YFP。可以用荧光显微镜检测这些 YFP 阳性细胞，通过流式细胞术进行定量，或使用荧光激活细胞分选收集。分离的 YFP 阳性细胞可以进行监测，以确定融合细胞的命运，特别是用于检测融合细胞的生长、转化和染色体数目的变化速度。
