Kopen G C, Prockop D J, Phinney D G
Center for Gene Therapy, MCP Hahnemann University, Philadelphia, Pennsylvania, USA.
J Histochem Cytochem. 1999 Jul;47(7):965-8. doi: 10.1177/002215549904700714.
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.
我们在此概述一种用于在经乙二醇甲基丙烯酸酯（GMA）处理的小鼠组织中进行β-葡萄糖醛酸酶高分辨率原位定位的方案。首先用5-溴-4-氯-3-吲哚基-β-D-葡萄糖醛酸（X-葡糖）对小鼠组织进行染色，然后在GMA中进行组织学处理。在X-葡糖中过夜孵育后蓝色靛蓝反应产物的保留使得通过明场显微镜能够对β-葡萄糖醛酸酶活性进行高分辨率定位。在暗场照明下，X-葡糖信号增强，即使在酶活性水平较低的细胞中也能检测到。该技术首次提供了一种更灵敏的酶组织化学方法来检测动物组织中的β-葡萄糖醛酸酶活性，同时也提供了在高倍放大下检查表达的机会。
