Immunohistochemical detection of beta-galactosidase or green fluorescent protein on tissue sections.
With the recent advances in mouse genetics, it is now possible to mark specific cell types genetically in vivo and to study the fate of cells during development and adulthood. Cells are labeled and followed in vivo through the stable expression of reporter genes in particular cell types. The two most commonly used reporter genes are LacZ, which encodes the enzyme beta-galactosidase (beta-gal), and green fluorescent protein (GFP). beta-Gal expression can be detected enzymatically, using 5-bromo-4-chloro-3-indolyl-beta-D-galactopyranoside (X-gal) as a substrate, and GFP can be directly visualized by fluorescence microscopy. However, with single detection of beta-gal or GFP, it is often impossible to determine whether expression of the reporter protein is restricted to a particular cell type. To ascertain the identity of individual cells within a multicellular tissue, beta-gal or GFP proteins must be visualized in conjunction with additional cellular markers. For such experiments, specific antibodies raised against beta-gal or GFP can be used in coimmunofluorescence analyses. Such double-staining analyses on tissue sections are a powerful tool to study transgene expression or to trace cells in multicellular tissues.
Seymour
Philip A
PA
Department of Developmental and Cell Biology, University of California, Irvine, USA.
Sander
Maike
M
D000818:Animals; D005455:Fluorescent Antibody Technique; D005697:Galactosides / Q000378:metabolism; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins / Q000032:analysis* / Q000276:immunology; D007150:Immunohistochemistry / Q000379:methods*; D007211:Indoles / Q000378:metabolism; D051379:Mice; D013194:Staining and Labeling; D001616:beta-Galactosidase / Q000032:analysis* / Q000276:immunology
D005697:Galactosides; D007211:Indoles; D049452:Green Fluorescent Proteins; D001616:beta-Galactosidase; C044888:5-bromo-4-chloro-3-indolyl beta-galactoside