| In vitro Fluorescent Matrix Degradation Assay for Entamoeba histolytica
|
|
Author:
Date:
2016-03-20
[Abstract] Fluorescent matrix degradation assay is a popular and widely used assay in the field of invadopodium biology (Artym et al., 2009). Matrix remodeling and degradation can be observed under both physiological and pathological conditions. Cancer cells extensively remodel and degrade the underlying matrix by employing actin-rich protrusive structures called invadosomes. Similar structures are formed by the protozoan parasite Entamoeba histolytica (E. histolytica), upon coming in contact with fibronectin, a major component of the host (extracellular matrix) ECM. Here, we describe a similar assay to measure matrix degradation by Entamoeba histolytica.
[摘要] 荧光基质降解测定是在invadopodium生物学领域中广泛使用的测定（Artym等人，2009）。 可以在生理和病理条件下观察基质重塑和降解。 癌细胞通过采用称为invadosomes的富含肌动蛋白的突出结构广泛地重塑和降解基础矩阵。 在与纤维连接蛋白（宿主（细胞外基质）ECM的主要组分）接触时，由原生动物寄生虫（即溶组织内阿米巴）（ histolytica ）形成类似的结构。 在这里，我们描述了通过溶组织内阿米巴测量基质降解的类似测定法。
|
|
|
| Pectin Nanostructure Visualization by Atomic Force Microscopy
|
|
Author:
Date:
2015-09-20
[Abstract] Pectins, complex polysaccharides rich in galacturonic acid, are a major component of primary plant cell walls. These macromolecules regulate cell wall porosity and intercellular adhesion, being important in the control of cell expansion and differentiation through their effect on the rheological properties of the cell wall. In fruits, pectin disassembly during ripening is one the main event leading to textural changes and softening. Changes in pectic polymer size, composition and structure have been studied by conventional techniques, most of them relying on bulk analysis of a population of polysaccharides but studies of detailed structure of isolated polymer chains are scarce (Paniagua et al., 2014). Atomic force microscopy (AFM) is a versatile and powerful technique able to ...
[摘要] 果胶，富含半乳糖醛酸的复合多糖是主要植物细胞壁的主要成分。这些大分子调节细胞壁孔隙度和细胞间粘附，在通过它们对细胞壁的流变性质的影响来控制细胞扩增和分化中是重要的。在果实中，果胶在成熟期间的拆卸是导致质地变化和软化的主要事件。已经通过常规技术研究了果胶聚合物尺寸，组成和结构的变化，其中大多数依赖于多糖群体的批量分析，但是对分离的聚合物链的详细结构的研究很少（Paniagua等人， >，2014）。原子力显微镜（AFM）是一种通用且强大的技术，能够分析力测量，以及以纳米尺度可视化生物样品的粗糙度（Morris等人，2010）。使用这种技术，最近的研究发现果胶纳米结构复杂性和几种果实的纹理和收获后行为之间的密切关系（Liu和Cheng，2011; Cybulska等人，2014; Pose等。，2015）。在这里，我们描述了AFM程序，以在地形学上可视化来自果实细胞壁提取物的果胶聚合物，其已经成功地用于草莓成熟的研究中（Pose等人，2012; ，2015）。因此，从AFM图像，可以在高放大率和最小的样品制备下分离孤立链（长度，高度和分支模式）的3D结构分析。 AFM基本原理和不同采样模式的完整描述在Morris等人（2010）中描述。
|
|
|
| Sandwich Enzyme-linked Immunosorbent Assay (ELISA) Analysis of Plant Cell Wall Glycan Connections
|
|
Author:
Date:
2014-04-20
[Abstract] Sandwich ELISA is a highly sensitive method that can be used to determine if two epitopes are part of the same macromolecule or supramolecular complex. In the case of plant cell wall glycans, it can reveal the existence of inter-polymers linkages, leading to better understanding of overall cell wall architectures. This development of a conventional sandwich ELISA protocol uses a carbohydrate-binding module (CBM), a small protein domain found in some carbohydrate catalysing or activating enzymes, and rat monoclonal antibodies (mAbs) which can be combined in the same ELISA plate without risk of cross reaction; the secondary anti-rat HRP antibody being only able to bind to the rat mAb and not the CBM. This protocol was developed and modified in the Prof. J. Paul Knox lab at the University of ...
[摘要] 夹心ELISA是高度灵敏的方法，其可以用于确定两个表位是否是相同大分子或超分子复合物的一部分。 在植物细胞壁聚糖的情况下，它可以揭示存在的聚合物之间的联系，导致更好地了解整体细胞壁结构。 常规夹心ELISA方案的这种发展使用碳水化合物结合模块（CBM），在一些碳水化合物催化或活化酶中发现的小蛋白结构域和大鼠单克隆抗体（mAb），其可以组合在相同的ELISA板中，没有风险 交叉反应; 次级抗大鼠HRP抗体仅能够结合大鼠mAb，而不能结合CBM。 该协议在利兹大学的J. Paul Knox实验室开发和修改。
|
|
|