Characterisation of two antibodies to oligomeric Aβ and their use in ELISAs on human brain tissue homogenates
Characterisation of two antibodies to oligomeric Aβ and their use in ELISAs on human brain tissue homogenates
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DOI：
10.1016/j.jneumeth.2008.09.002
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发表时间：
2009-01
影响因子：
3
通讯作者：
Z. V. Helmond;K. Heesom;S. Love
中科院分区：
文献类型：
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作者：
Z. V. Helmond;K. Heesom;S. Love
寡聚体形式的Aβ被认为是阿尔茨海默病（AD）中该肽的主要毒性种类。尽管已经报道了寡聚体特异性抗体的表征，但这些抗体尚未成功地纳入酶联免疫吸附试验（ELISA）中，并且脑组织中寡聚体Aβ水平的测量仍然存在问题。我们检测了两种单克隆抗体7A 1a和1G 5对合成的Aβ1- 42寡聚体和人脑匀浆中寡聚体Aβ1- 42的特异性，以及这两种抗体用于夹心ELISA测定寡聚体Aβ1- 42的实用性。发现两种抗体都能识别一系列Aβ1- 42的合成寡聚体，但在Western印迹中与一种34 kDa的蛋白质发生交叉反应，该蛋白质经二维凝胶电泳和质谱分析显示为原肌球蛋白。然而，通过将7A 1a和1G 5与Aβ1- 42捕获抗体结合使用，我们能够通过ELISA特异性地检测和测量脑匀浆中寡聚体Aβ1- 42的水平。建立一种简单的ELISA检测寡聚体Aβ的方法，有助于进一步研究Aβ寡聚体在AD中的作用。
Oligomeric forms of Aβ are believed to be the major toxic species of this peptide in Alzheimer's disease (AD). Although the characterisation of oligomer-specific antibodies has been reported, these have not been successfully incorporated into an enzyme-linked immunosorbent assay (ELISA), and measurement of the levels of oligomeric Aβ in brain tissue has remained problematic. We have examined the specificity of two monoclonal antibodies, 7A1a and 1G5, for synthetic oligomers of Aβ1–42and for oligomeric Aβ1–42in human brain homogenates, and the utility of these two antibodies for measuring oligomeric Aβ1–42by sandwich ELISA. Both antibodies were found to recognise a range of synthetic oligomers of Aβ1–42but to cross-react in Western blots with a 34kDa protein, shown by two-dimensional gel electrophoresis and mass spectrometry to be tropomyosin. However, by using 7A1a and 1G5 in combination with an Aβ1–42capture antibody, we were able specifically to detect and to measure the levels of oligomeric Aβ1–42in brain homogenates by ELISA. The development of a simple ELISA for measurement of oligomeric Aβ should facilitate further studies of the role of oligomeric species of Aβ in AD.