Optimized Immunohistochemical Detection of Rat ESR2 Proteins Using the Specific Anti-ESR2 Monoclonal Antibody PPZ0506
Optimized Immunohistochemical Detection of Rat ESR2 Proteins Using the Specific Anti-ESR2 Monoclonal Antibody PPZ0506
复制标题
使用特定的抗ESR2单克隆抗体PPZ0506对大鼠ESR2蛋白进行优化的免疫组织化学检测
DOI：
10.1210/jendso/bvab048.1656
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发表时间：
2021-05-03
影响因子：
4.1
通讯作者：
Ozawa H
中科院分区：
文献类型：
--
作者：
Hattori Y;Ishii H;Higo S;Otsuka M;Kanaya M;Matsumoto K;Ozawa M;Ozawa H
背景资料：对ESR 2的研究，也被称为雌激素受体β（ERβ），是由于使用未经充分验证的抗体而导致数据失真的一个臭名昭著的例子。尽管缺乏可靠的抗ESR 2特异性抗体严重阻碍了ESR 2研究的推广，但2017年鉴定出了一种特异性抗人ESR 2单克隆抗体（PPZ 0506）[1]。我们之前的研究证实了其对啮齿动物ESR 2蛋白的交叉反应性和特异性，从而能够阐明啮齿动物中的真实ESR 2分布[2]。目的：确定PPZ 0506免疫组化检测大鼠ESR 2蛋白的最佳条件。<Method>对石蜡包埋和冷冻卵巢切片的几种染色条件进行了评价，并在最佳条件下分析了大鼠ESR 2蛋白的分布。结果：PPZ 0506免疫组化染色需要适当的抗原修复和抗体稀释。随后在优化条件下在多个组织中进行的免疫组织化学分析显示，大鼠ESR 2蛋白的表达比先前假设的更局部化。我们使用经过充分验证的抗体对大鼠ESR 2蛋白进行了优化的免疫组织化学检测，揭示了它们在有限组织和细胞类型中的分布。结论：我们的研究结果表明，以前的免疫组织化学研究使用不充分验证的抗体对ESR 2蛋白高估了他们的分布概况。我们希望我们使用PPZ 0506抗体的优化免疫组织化学可以解决ESR 2研究中的冲突问题。参考文献：1. Andersson S，et al. Nat Commun 15;8：15840（2017）2.石井H等人Int J Mol Sci 20（24）：6312（2019）
Background: Research on ESR2, also known as estrogen receptor β (ERβ), is a notorious example of data distortion due to the use of inadequately validated antibodies. Although the absence of reliable specific antibodies against ESR2 has severely hindered the promotion of ESR2 research, a specific anti-human ESR2 monoclonal antibody (PPZ0506) was identified in 2017 [1]. Our previous study confirmed its cross-reactivity and specificity against rodent ESR2 proteins, enabling the elucidation of the true ESR2 distribution in rodents [2]. Objective: We aimed to determine the optimized conditions for immunohistochemical detection of rat ESR2 proteins using PPZ0506. <Method> Several staining conditions using paraffin-embedded and frozen ovary sections were evaluated, and the distribution of rat ESR2 proteins was analyzed under optimal conditions. Result: Immunohistochemical staining with PPZ0506 required appropriate antigen retrieval and antibody dilution. Subsequent immunohistochemical analysis in multiple tissues under optimized conditions revealed that rat ESR2 proteins are expressed in a more localized manner than previously assumed. Our optimized immunohistochemical detection of rat ESR2 proteins, using a well-validated antibody, revealed their distribution in limited tissues and cell types. Conclusion: Our results suggest that previous immunohistochemical studies using inadequately validated antibodies against ESR2 proteins overestimated their distribution profiles. We expect that our optimized immunohistochemistry using the PPZ0506 antibody may solve conflicting problems in ESR2 research. References: 1. Andersson S, et al. Nat Commun 15;8:15840 (2017) 2. Ishii H, et al. Int J Mol Sci 20(24):6312 (2019)