Development and characterization of monoclonal antibodies against human aryl hydrocarbon receptor

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• 作者：Wenjing Tian¹ ; Xinhui Pei¹ ; Heidi Qunhui Xie¹ ; ^{qhxie@rcees.ac.cn" class="auth_mail" title="E-mail the corresponding author} ; Sherry Li Xu¹ ; Jijing Tian¹ ; Qin Hu¹ ; Haiming Xu¹ ; Yangsheng Chen¹ ; Hualing Fu¹ ; Zhengyu Cao² ; ^{zycao1999@hotmail.com" class="auth_mail" title="E-mail the corresponding author} ; Bin Zhao¹ ; ^{binzhao@rcees.ac.cn" class="auth_mail" title="E-mail the corresponding author}
• 关键词：Aryl hydrocarbon receptor ; Monoclonal antibody ; Western blot ; Immunoprecipitation ; Indirect immunofluorescence assay
• 刊名：Journal of Environmental Sciences
• 出版年：2016
• 出版时间：January 2016
• 年：2016
• 卷：39
• 期：Complete
• 页码：165-174
• 全文大小：825 K

文摘

Aryl hydrocarbon receptor (AhR), a ligand-dependent nuclear receptor, is involved in a diverse spectrum of biological and toxicological effects. Due to the lack of three dimensional (3D) crystal or nuclear magnetic resonance structure, the mechanisms of these complex effects of AhR remain to be unclear. Also, commercial monoclonal antibodies (mAbs) against human AhR protein (hAhR), as alternative immunological tools, are very limited. Thus, in order to provide more tools for further studies on hAhR, we prepared two mAbs (1D6 and 4A6) against hAhR. The two newly generated mAbs specifically bound to amino acids 484–508 (located in transcription activation domain) and amino acids 201–215 (located in Per-ARNT-Sim domain) of hAhR, respectively. These epitopes were new as compared with those of commercial mAbs. The mAbs were also characterized by enzyme-linked immunosorbent assay, western blot, immunoprecipitation and indirect immunofluorescence assay in different cell lines. The results showed that the two mAbs could recognize the linearized AhRs in six different human cell lines and a rat hepatoma cell line, as well as the hAhR with native conformations. We concluded that the newly generated mAbs could be employed in AhR-based bioassays for analysis of environmental contaminants, and held great potential for further revealing the spatial structure of AhR and its biological functions in future studies.