Evaluation of seven oestrogen receptor β antibodies for immunohistochemistry, western blotting, and flow cytometry in human breast tissue
Evaluation of seven oestrogen receptor β antibodies for immunohistochemistry, western blotting, and flow cytometry in human breast tissue
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DOI：
10.1002/path.1077
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发表时间：
2002-06-01
影响因子：
7.3
通讯作者：
Speirs, V
中科院分区：
文献类型：
--
作者：
Skliris, GP;Parkes, AT;Speirs, V
存在两种雌激素受体，ER α和ER β。虽然对ER α了解很多，但ER β的作用仍然不确定，特别是在蛋白质水平。本研究的目的是确定针对ER β不同结构域的7种ER β抗体（14 C8、8D 5、PA 1313、PPG 5/10、N19、9.88和D 7 N）在三种常用实验室应用（即免疫组织化学、蛋白质印迹和流式细胞术）中的效用，使用人乳腺材料。对于冷冻材料的免疫组织化学分析，PA 1313和D 7 N比14 C8、8D 5和PPG 5/10产生更强和更特异的信号。在石蜡切片中，14 C8，紧随其后的PPG 5/10，给出了迄今为止最上级的核免疫反应性，与测试的其他抗体相比。通常，流式细胞术结果反映了石蜡切片的免疫组织化学数据，其中抗体等级为14 C8> 8D 5大于或等于派-313 > PPG 5/10 > D 7 N。对于蛋白质印迹，8135和D 7 N产生最强和最一致的条带，与其他人一起观察到较弱的条带。结论：ER β蛋白可以用特异性抗体检测。然而，这些抗体的特异性和应用之间存在相当大的差异，突出了在选择用于特定实验室技术的抗体时需要仔细优化的事实。版权所有（C）2002约翰威利父子有限公司
Two oestrogen receptors, ERalpha and ERbeta, exist. While much is known about ERalpha, the role of ERbeta is still undefined, especially at the protein level. The aim of this study was to determine the utility of seven ERbeta antibodies (14C8, 8D5, PA1313, PPG5/10, N19, 9.88, and D7N) raised against different domains of ERbeta in three commonly used laboratory applications, namely, immunohistochemistry, western blot, and flow cytometry, using human breast material. For immunohistochemical analysis of frozen material, PA1313 and D7N gave stronger and more specific signals than 14C8, 8D5, and PPG5/10. In paraffin sections, 14C8, closely followed by PPG5/10, gave by far the most superior nuclear immunoreactivity, compared with the other antibodies tested. In general, flow, cytometry results mirrored the immunohistochemistry data for paraffin sections, with antibodies ranked 14C8 > 8D5 greater than or equal to PAI-313 > PPG5/10 > D7N. For western blotting, 8135 and D7N yielded the strongest and most consistent bands, with weaker bands seen with the others. It is concluded that ERbeta protein can be detected using specific antibodies. However, there is onsiderable variation between the specificity and application of these antibodies, highlighting the fact that careful optimization is required when selecting an antibody for use in a particular laboratory technique. Copyright (C) 2002 John Wiley Sons, Ltd.