Detailed specificity analysis of antibodies binding to modified histone tails with peptide arrays
Detailed specificity analysis of antibodies binding to modified histone tails with peptide arrays
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DOI：
10.4161/epi.6.2.13837
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发表时间：
2011-02-01
期刊：
影响因子：
3.7
通讯作者：
Jeltsch, Albert
中科院分区：
文献类型：
--
作者：
Bock, Ina;Dhayalan, Arunkumar;Jeltsch, Albert
组蛋白尾部翻译后修饰（PTM）对染色质结构有很大的影响，在表观遗传过程中也起着重要作用。针对修饰的组蛋白尾部的抗体是染色质生物学和分子表观遗传学的中心研究试剂。我们应用Celluspots肽阵列对来自不同供应商的36种针对修饰的组蛋白尾部的商业抗体进行特异性分析。该阵列包含来自组蛋白N-末端尾部的8个不同区域的384个肽，即H3 1-19、7-26、16-35和26-45、H4 1-19和11-30、H2 A 1-19和H2 B 1-19，其特征在于以许多不同组合的59个翻译后修饰。使用各种控制，我们记录的方法的可靠性。我们的分析揭示了先前未记录的测试抗体的特异性特征的细节。大多数抗体与PTM结合良好，但有些抗体失败了。此外，一些抗体显示出高交叉反应性，并且大多数抗体被接近主要PTM的特异性额外PTM抑制。此外，针对相同修饰的抗体的特异性特征有时非常不同。表观遗传学研究中使用的抗体的特异性是一个重要问题。我们提供了36个广泛使用的商业组蛋白尾PTM抗体的抗体特异性概况的目录。更好地了解抗体的特异性特征将使研究人员能够在生物学研究中实施必要的对照实验，并允许使用这些抗体对生物学实验进行更可靠的解释。
Chromatin structure is greatly influenced by histone tail post-translational modifications (PTM), which also play a central role in epigenetic processes. Antibodies against modified histone tails are central research reagents in chromatin biology and molecular epigenetics. We applied Celluspots peptide arrays for the specificity analysis of 36 commercial antibodies from different suppliers which are directed towards modified histone tails. The arrays contained 384 peptides from eight different regions of the N-terminal tails of histones, viz. H3 1-19, 7-26, 16-35 and 26-45, H4 1-19 and 11-30, H2A 1-19 and H2B 1-19, featuring 59 post-translational modifications in many different combinations. Using various controls we document the reliability of the method. Our analysis revealed previously undocumented details in the specificity profiles of the tested antibodies. Most of the antibodies bound well to the PTM they have been raised for, but some failed. In addition, some antibodies showed high cross-reactivity and most antibodies were inhibited by specific additional PTMs close to the primary one. Furthermore, specificity profiles for antibodies directed toward the same modification sometimes were very different. The specificity of antibodies used in epigenetic research is an important issue. We provide a catalog of antibody specificity profiles for 36 widely used commercial histone tail PTM antibodies. Better knowledge about the specificity profiles of antibodies will enable researchers to implement necessary control experiments in biological studies and allow more reliable interpretation of biological experiments using these antibodies.