Enhanced in situ detection of β-glucuronidase activity in murine tissue

Abstract

We outline here a protocol for high-resolution in situ localization of β-glucuronidase in murine tissues processed in glycol methacrylate (GMA). Murine tissues were first stained with 5-bromo-4-chloro-3-indolyl-β-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 β-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 β-glucuronidase activity in animal tissues and also the opportunity to examine expression at high magnification.