Biotechnology Research Group, Fundamental Research Department, Fujirebio Inc., Tokyo 192-0031, Japan.
Biochem Biophys Res Commun. 2010 Apr 9;394(3):685-90. doi: 10.1016/j.bbrc.2010.03.052. Epub 2010 Mar 11.
The precise regulation of epidermal growth factor receptor (EGFR) is crucial for its function in cellular growth control. Although many antibodies against EGFR have been developed and used to analyze its regulation and function, it is not yet easy to analyze activated EGFR specifically. Activated EGFR has been mainly detected by its phosphorylation state using anti-phospho-EGFR and anti-phosphotyrosine antibodies. In the present study, we have established novel monoclonal antibodies which recognize the activated EGFR independently of its phosphorylation. Our antibodies detected active state of EGFR in immunoprecipitation and immunofluorescence, by recognizing the epitopes which are exposed through the conformational change induced by ligand-binding. Furthermore, we found that our antibodies preferentially detected the conformation of constitutively active EGFR mutants found in lung cancer cell lines. These results indicate that our antibodies may become novel research and diagnostic tools for detecting and analyzing the conformation of active EGFR in various cells and tissues.
表皮生长因子受体 (EGFR) 的精确调节对于其在细胞生长控制中的功能至关重要。尽管已经开发并使用了许多针对 EGFR 的抗体来分析其调节和功能，但特异性分析激活的 EGFR 并不容易。激活的 EGFR 主要通过使用抗磷酸化-EGFR 和抗磷酸酪氨酸抗体检测其磷酸化状态来检测。在本研究中，我们建立了新型单克隆抗体，这些抗体可识别配体结合诱导构象变化后暴露的表位，而无需依赖 EGFR 的磷酸化来识别激活的 EGFR。我们的抗体通过识别配体结合诱导的构象变化后暴露的表位，在免疫沉淀和免疫荧光中检测到 EGFR 的活性状态。此外，我们发现我们的抗体优先检测到在肺癌细胞系中发现的组成性激活 EGFR 突变体的构象。这些结果表明，我们的抗体可能成为检测和分析各种细胞和组织中活性 EGFR 构象的新型研究和诊断工具。
