Immunoblotting with Peptide Antibodies: Differential Immunoreactivities Caused by Certain Amino Acid Substitutions in a Short Peptide and Possible Effects of Differential Refolding of the Peptide on a Nitrocellulose or PVDF Membrane
Immunoblotting with Peptide Antibodies: Differential Immunoreactivities Caused by Certain Amino Acid Substitutions in a Short Peptide and Possible Effects of Differential Refolding of the Peptide on a Nitrocellulose or PVDF Membrane
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使用肽抗体进行免疫印迹：短肽中某些氨基酸取代引起的差异免疫反应性以及肽在硝化纤维或 PVDF 膜上差异重折叠的可能影响
DOI：
10.1007/978-1-4939-2999-3_26
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
Yasuo Shinohara
中科院分区：
文献类型：
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作者：
Takenori Yamamoto;Taisuke Matsuo;Atsushi Yamamoto;Ryohei Yamagoshi;Kazuto Ohkura;Masatoshi Kataoka;Yasuo Shinohara
使用抗体的免疫检测，例如Western blotting，通常用于测量蛋白质混合物中某种蛋白质的量。为了有效地解释免疫检测结果，必须严格注意影响抗体免疫反应性的因素。我们在这里描述了一步一步的过程，以证明在肽中取代某些氨基酸可以引起其与固定肽中针对表位标签的抗体的免疫反应性的显着差异。膜上肽的重新折叠在不同程度上掩盖了抗原表位，这可能是它们与抗体产生不同免疫反应的原因。本章的结果提示我们需要仔细解释涉及免疫检测的实验结果。
Immunodetection using antibodies, e.g., Western blotting, is generally utilized to measure the amount of a certain protein in a protein mixture. For valid interpretation of results observed by immunodetection, strict attention must be paid to the factors affecting the immunoreactivities of the antibodies. We here describe the step-by-step procedures to demonstrate that substitution of certain amino acids in a peptide can cause remarkable differences in its immunoreactivity with antibodies against epitope tags in the immobilized peptide. Refolding of the peptide on the membrane in a way that masks the epitope to different degrees was the possible reason for their distinct immunoreactivities with the antibodies. The results in this chapter suggest that we need to interpret carefully the experimental results involving immunodetection.