Generation and characterization of antibodies against arginine-derived advanced glycation endproducts
Generation and characterization of antibodies against arginine-derived advanced glycation endproducts
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DOI：
10.1016/j.bmcl.2015.06.013
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发表时间：
2015-11-01
影响因子：
2.7
通讯作者：
Spiegel, David A.
中科院分区：
文献类型：
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作者：
Wang, Tina;Streeter, Matthew D.;Spiegel, David A.
虽然抗体试剂已被广泛用于研究晚期糖基化终产物（AGEs），但这些材料是使用免疫原的复杂混合物生产的。因此，它们的表位特异性仍然未知。在这里，我们已经产生了第一个抗体，能够识别的三个异构体的甲基乙二醛氢咪唑酮（MG-H）通过使用化学合成，以创建同质免疫原。此外，我们通过采用合成MG-H抗原实施直接ELISA方案，彻底表征了我们的抗体和两种现有单克隆抗体的表位特异性。最后，我们采用已报道的抗MG-H抗体，使用免疫荧光显微镜检测细胞系统中的MG-H。这些研究表明，抗MG-H1和抗MG-H3染色集中在细胞核内，而抗MG-H2仅提供最小的信号。这些观察结果与报道的MG-Hs形成偏好一致，并可能表明非酶促翻译后修饰的新核靶点。本文报道的抗体试剂，以及用于其创建的策略，可能被证明是有用的AGEs在生物系统中的免疫化学研究。(C)2015爱思唯尔有限公司版权所有。
Although antibodies reagents have been widely employed for studying advanced glycation end-products (AGEs), these materials have been produced using complex mixtures of immunogens. Consequently, their epitope specificity remains unknown. Here we have generated the first antibodies capable of recognizing each of the three isomers of the methylglyoxal hydroimidazolones (MG-Hs) by using chemical synthesis to create homogenous immunogens. Furthermore, we have thoroughly characterized the epitope specificity of both our antibodies and that of two existing monoclonals by implementing a direct ELISA protocol employing synthetic MG-H antigens. Finally, we employed the reported anti-MG-H antibodies to the detection of MG-Hs in cellular systems using immunofluorescence microscopy. These studies have demonstrated that anti-MG-H1 and anti-MG-H3 staining is concentrated within the nucleus, while anti-MG-H2 affords only minimal signal. These observations are consistent with reported formation preferences for MG-Hs, and may suggest novel nuclear targets for non-enzymatic posttranslational modification. The antibody reagents reported herein, as well as the strategy employed for their creation, are likely to prove useful for the immunochemical study of AGEs in biological systems. (C) 2015 Elsevier Ltd. All rights reserved.