Enhanced in situ detection of beta-glucuronidase activity in murine tissue.
We outline here a protocol for high-resolution in situ localization of beta-glucuronidase in murine tissues processed in glycol methacrylate (GMA). Murine tissues were first stained with 5-bromo-4-chloro-3-indolyl-beta-d-glucuronic acid (x-gluc), followed by histological processing in GMA. Retention of the blue indigo reaction product after overnight incubations in x-gluc allowed high-resolution localization of beta-glucuronidase activity by brightfield microscopy. When illuminated under darkfield, the x-gluc signal was enhanced, permitting detection even in cells with low-level enzyme activity. This technique offers for the first time a more sensitive enzyme histochemical method of detecting beta-glucuronidase activity in animal tissues and also the opportunity to examine expression at high magni-fication.
Kopen
G C
GC
Center for Gene Therapy, MCP Hahnemann University, Philadelphia, Pennsylvania, USA.
Prockop
D J
DJ
Phinney
D G
DG
D000818:Animals; D005965:Glucuronates / Q000737:chemistry; D020723:Glucuronic Acid; D005966:Glucuronidase / Q000032:analysis*; D007124:Immunoenzyme Techniques; D008099:Liver / Q000201:enzymology; D008689:Methacrylates; D051379:Mice; D013154:Spleen / Q000201:enzymology
D005965:Glucuronates; D008689:Methacrylates; C005044:hydroxyethyl methacrylate; D020723:Glucuronic Acid; D005966:Glucuronidase