Chapter 19: Flow Cytometric Analysis and FACS Sorting of Cells Based on GFP Accumulation

Abstract

Publisher Summary The use of green fluorescent proteins (GFP) as a reporter in flow cytometry and cell sorting is in its infancy. A major benefit provided by GFP is that it can be measured noninvasively. Alterations to the primary structure of the molecule have provided variants that are readily detectable in both prokaryotic and eukaryotic cells and within subcellular organelles. Further spectral variants that can be combined for multiparametric analyses will find considerable future utility in flow cytometry. In animal cell systems, GFP is particularly suited as an indicator of viral infection. When expressed in viruses, either as a gene fusion or when substituted for a nonessential viral gene, GFP fluorescence provides a sensitive and accurate measure of the proportions of infected cells and the progress of viral infection. Coupling this approach for viral detection with concerted mutagenesis of virus provides a screen for the roles of the various viral components within the infection process. Flow cytometric detection of GFP also finds practical application in gene therapy. By marking the viral vectors containing therapeutic gene products with GFP, the members expressing these products within a population of cells can be identified, selected, and used exclusively for the reconstitution of organisms.