Tai Phillip W L, Smith Catherine L, Angello John C, Hauschka Stephen D
Department of Biochemistry, University of Washington, Seattle, WA, USA.
Methods Mol Biol. 2012;798:445-59. doi: 10.1007/978-1-61779-343-1_26.
β-galactosidase (β-gal) is among the most frequently used markers for studying a wide variety of biological mechanisms, e.g., gene expression, cell migration, stem cell conversion to different cell types, and gene silencing. Many of these studies require the histochemical detection of relative β-gal levels in tissue cross-sections mounted onto glass slides and visualized by microscopy. This is particularly useful for the analysis of promoter activity in skeletal muscle tissue since the β-gal levels can vary dramatically between different anatomical muscles and myofiber types. The differences in promoter activity can be due to a myofiber's developmental history, innervation, response to normal or experimental physiological signals, and its disease state. It is thus important to identify the individual fiber types within muscle cross-sections and to correlate these with transgene expression signals. Here, we provide a detailed description of how to process and analyze muscle tissues to determine the fiber-type composition and β-gal transgene expression within cryosections.
β-半乳糖苷酶（β-gal）是研究多种生物学机制时最常用的标记物之一，例如基因表达、细胞迁移、干细胞向不同细胞类型的转化以及基因沉默。这些研究中的许多都需要对安装在载玻片上并通过显微镜观察的组织切片中的相对β-gal水平进行组织化学检测。这对于分析骨骼肌组织中的启动子活性特别有用，因为不同解剖学肌肉和肌纤维类型之间的β-gal水平可能有很大差异。启动子活性的差异可能归因于肌纤维的发育历史、神经支配、对正常或实验性生理信号的反应及其疾病状态。因此，识别肌肉切片中的单个纤维类型并将其与转基因表达信号相关联非常重要。在这里，我们详细描述了如何处理和分析肌肉组织，以确定冷冻切片内的纤维类型组成和β-gal转基因表达。
