BRCA1 Antibodies Matter.
BRCA1 Antibodies Matter.
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DOI：
10.7150/ijbs.63115
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发表时间：
2021
影响因子：
9.2
通讯作者：
Hu Y
中科院分区：
文献类型：
--
作者：
Yang J;Qi L;Chiang HC;Yuan B;Li R;Hu Y
乳腺癌易感基因 1 (BRCA1) 编码一种肿瘤抑制基因，该基因在家族性乳腺癌和卵巢癌患者中经常发生突变。 BRCA1 在多个重要的细胞过程中发挥作用，包括 DNA 损伤修复、细胞周期检查点激活、蛋白质泛素化、染色质重塑、转录调节以及 R 环形成和细胞凋亡。在过去的三十年里，大量的 BRCA1 抗体已经产生并投入商业使用，然而，许多商业抗体的特征描述很差，并且在广泛使用时导致数据不可靠。为了寻找可靠且特异的 BRCA1 抗体 (Abs)，特别是识别小鼠 BRCA1 的抗体，我们对许多市售抗 BRCA1 抗体进行了严格验证，在一组验证应用中使用适当的对照，包括蛋白质印迹 (WB)、免疫沉淀 (IP)、免疫沉淀-质谱 (IP-MS)、染色质免疫沉淀 (ChIP) 和免疫荧光 (IF)。此外，我们通过使用来自 Brca1+/+ 和 Brca1Δ11/Δ11 小鼠的睾丸组织和小鼠胚胎成纤维细胞 (MEF) 评估了这些抗体检测小鼠 BRCA1 蛋白的特异性。我们发现 Ab1、D-9、07-434（用于识别人类 BRCA1）和 287.17、440621、BR-64（用于识别小鼠 BRCA1）在所示测定中具有特异性和高质量性能。我们在这里分享这些结果的目的是帮助社区应对与抗 BRCA1 抗体特异性和重现性相关的常见挑战，并希望更好地了解细胞和组织水平上的 BRCA1 功能。
Breast cancer susceptibility gene 1 (BRCA1) encodes a tumor suppressor that is frequently mutated in familial breast and ovarian cancer patients. BRCA1 functions in multiple important cellular processes including DNA damage repair, cell cycle checkpoint activation, protein ubiquitination, chromatin remodeling, transcriptional regulation, as well as R-loop formation and apoptosis. A large number of BRCA1 antibodies have been generated and become commercially available over the past three decades, however, many commercial antibodies are poorly characterized and, when widely used, led to unreliable data. In search of reliable and specific BRCA1 antibodies (Abs), particularly antibodies recognizing mouse BRCA1, we performed a rigorous validation of a number of commercially available anti-BRCA1 antibodies, using proper controls in a panel of validation applications, including Western blot (WB), immunoprecipitation (IP), immunoprecipitation-mass spectrometry (IP-MS), chromatin immunoprecipitation (ChIP) and immunofluorescence (IF). Furthermore, we assessed the specificity of these antibodies to detect mouse BRCA1 protein through the use of testis tissue and mouse embryonic fibroblasts (MEFs) from Brca1+/+ and Brca1Δ11/Δ11 mice. We find that Ab1, D-9, 07-434 (for recognizing human BRCA1) and 287.17, 440621, BR-64 (for recognizing mouse BRCA1) are specific with high quality performance in the indicated assays. We share these results here with the goal of helping the community combat the common challenges associated with anti-BRCA1 antibody specificity and reproducibility and, hopefully, better understanding BRCA1 functions at cellular and tissue levels.
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影响因子：
5.8
作者：
Holmstrom, Sam R.;Wijayatunge, Ranjula;Ross, Theodora S.
通讯作者：
Ross, Theodora S.
影响因子：
11.2
作者：
De Siervi A;De Luca P;Byun JS;Di LJ;Fufa T;Haggerty CM;Vazquez E;Moiola C;Longo DL;Gardner K
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作者：
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通讯作者：
Komatsu Y
影响因子：
56.9
作者：
Cortez, D;Wang, Y;Elledge, SJ
通讯作者：
Elledge, SJ