Fluorescence–activated cell sorting (FACS) was invented to sort a heterogeneous mixture of cells into different homogenous subpopulations of interest based upon the specific light scattering and fluorescent characteristics of each cell. In a complex cell mixture, the different homogenous subpopulations may have different antigenic and other markers on their surface. These markers can be tagged by the means of fluorescent labels which can be detected by laser and light detectors of fluorescence-activated cell sorting. It provides rapid, accurate and high throughput cell analysis and has been applied to examine diverse biological processes such as cell cycle, cell proliferation, cell viability, cell phenotyping, cell signaling, micronucleus test, intracellular cytokine secretion.
Figure 1. The basic principle of FACS
Fluorescence–activated cell sorting determine cells automatically based either on cellular properties or by fluorescent labeling. The ability to sort cells based on physical characteristic and their fluorescent label signatures enables to isolate well-defined subpopulations of cells in more effective manner than other separation methods likely magnetic separation. The use of fluorescent colors facilitates to detect simultaneously different subpopulations of interest, it is time-effective and labor-saving. Green fluorescent protein (GFP) is used as a reporter gene. another complementary is monoclonal antibodies, which is highly specific for its target antigens and can readily be coupled with fluorescein, phycobiliproteins and other fluorochromes. Monoclonal antibodies widely used as FACS reagents enhanced the definition of hundreds of target antigens present on or in cells. FACS is also the most efficient method of cloning cells, especially when they are present in a extremely low frequency.
Figure 2. The experimental results of FACS
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