Knox J Paul
Faculty of Biological Sciences, Centre for Plant Sciences, University of Leeds, Leeds, United Kingdom.
Methods Enzymol. 2012;510:233-45. doi: 10.1016/B978-0-12-415931-0.00012-4.
Cellulose is generally found in the context of complex plant cell wall materials and mostly in association with other glycans. Cellulose-directed carbohydrate-binding modules (CBMs) can be readily adapted to a range of methods for the in situ imaging of cellulose structures within plant cell walls or other cellulose-based materials. Protocols for the preparation and selection of plant materials, their fixation and processing for preparation of sections for CBM labeling, and fluorescence imaging procedures are described. Approaches to direct methods in which CBMs are directly coupled to fluorophores and indirect methods in which staged incubations with secondary reagents are used for the fluorescence imaging of CBM binding to materials are discussed and presented.
纤维素通常存在于复杂的植物细胞壁材料中,并且大多与其他聚糖结合。纤维素导向的碳水化合物结合模块(CBMs)可以很容易地应用于一系列方法,用于对植物细胞壁或其他纤维素基材料中的纤维素结构进行原位成像。本文描述了植物材料的制备和选择、固定以及为CBM标记制备切片的处理方法,以及荧光成像程序。文中讨论并介绍了将CBMs直接与荧光团偶联的直接方法和使用二级试剂进行分步孵育以对CBM与材料的结合进行荧光成像的间接方法。