Facts about FACS

Thu, 04/18/2013 - 15:49

FACS (Fluorescence Activated Cell Sorting) was developed by Bonner, Sweet, Hulett, Herzenberg, and others to perform flow cytometry. Flow cytometry is a powerful analytical tool useful for the characterization or phenotypic identification of different populations of cells. Characteristics of individual particles or cells are rapidly measured as they pass through light emitted from a laser.  As the laser light is scattered by the cell, cell size and complexity or granularity can be estimated by forward scatter (FSC) and side scatter (SSC), respectively.  An amalgamation of both measurements is helpful in discriminating multiple cell types in a cohort of cell population.

Over the years, there has been an upsurge in additional measurable FACS parameters, spurred by enhanced availability of fluorescently labeled antibodies.  Fluorochrome-conjugated antibodies also known as fluorescent probes are traditionally used in targeting epitopes present on the cell surface (1).  Recently, optimization of permeabilization and fixation buffers has also facilitated measurement of intracellular antigens via flow cytometry.  By labeling specific combinations of antigens with fluorochrome-conjugated antibodies, researchers are able to identify populations of cells with much greater certainty.  At the most basic level, this information can be used to quantify the abundance of particular populations of cells in a given sample.  The data may also be used to obtain measurements describing the relative expression levels of several proteins within particular populations of cells to provide additional details regarding protein expression compared to other techniques that only measure proteins in heterogeneous samples.

Data analysis in flow cytometry is generally preformed using single and two-parameter histograms comparing the relative density of each antigen which facilitates immunophenotyping. Gating of the cells based upon fluorescence intensities of each antigen plotted on these charts allows the researcher to selectively examine cells of interest, while disregarding the other populations of cells or cellular debris. Several FACS protocols are available for the sample preparation contingent to the cell type, method of staining and analysis. Novus Biologicals is proud to be at the fore front specializing in flow cytometry reagents and offer the latest range of antibody markers along with accessory buffers for your research needs.

  1. PMID: 22939984

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