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Allows ultra-sensitive detection of β-galactosidase activity in live mammalian, yeast, bacterial, or plant cells. Especially useful for FACS analysis.
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One of the most common reporter genes used in molecular biology applications is the E.Coli lacZ
gene that codes for an active subunit of beta-galactosidase in vivo
. Since this enzyme is generally absent in normal mammalian cells, it can be detected at very low levels, and since the enzyme has a wide substrate specificity, monitoring lacZ expression (and therefore co-expressed genes or promoter efficiency) has become routine to the point of detection of as few as 5 copies of beta-galactosidase per cell.
Although chromogenic assays of beta-galactosidase activity (i.e. X-Gal) have use, the recent application of the fluorogenic substrate fluorescein di-β-D-galactopyranoside (FDG) combined with fluorescence microscopic analysis (confocal microscopy) analysis has been shown to be several orders of magnitude more sensitive. In addition, because of its improved detection limits, the FDG substrate has found extensive use in automated ELISA type assay systems .
This kit uses the beta-galactoside analog fluorescein di-β-D-Galactopyranoside (FDG) in a protocol that sensitively distinguishes lacZ+ vs. lacZ- cells. See also Product M0250 for more information and references.
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- Fiering S, Nolan G, Herzenberg LA. (1988) "Inhibitors of endogenous mammalian beta-Galactosidase (B-Gal) or transduced E. coli B-Gal improve the sensitivity and versatility of the FACS-FDG fluorogenic Assay for E. coli lacZ expression in viable mammalian cells." Cytometry (Suppl 2) 1.
- Guo W, Wu H, (2008) "Detection of lacZ expression by FACS-Gal analysis" Nature Protocols DOI: 10.1038/nprot.2008.163 http://www.natureprotocols.com/2008/08/06/detection_of_lacz_expression_b.php