Diermeier-Daucher Simone, Brockhoff Gero
Institute of Pathology, University of Regensburg, Regensburg, Germany.
Curr Protoc Cytom. 2008 Jul;Chapter 12:Unit12.14. doi: 10.1002/0471142956.cy1214s45.
The homologous and heterologous interaction of members of the epidermal growth factor (EGF)-related receptor tyrosine kinase (RTK) family (ErbB or HER family receptors) upon ligand binding is the initial key event in signal transduction by these receptors. In addition to the availability of their respective ligands, the relative expression level of the four ErbB receptors triggers receptor cross-activation, which determines signal diversification and the cells' biological response. However, the function of ErbB receptors and their ligands appears highly complex, and its impact on cell growth and proliferation of normal and tumor cells is incompletely understood. Flow cytometric fluorescence resonance energy transfer (FRET) measurements facilitate the quantitative analysis of receptor interaction. This unit delineates the cell-by-cell analysis of epidermal growth factor receptor (EGFR, ErbB1, HER1) and ErbB2 (HER2) receptor interaction in ErbB2-overexpressing BT474 and SK-BR-3 breast cancer cell lines, using a dual-laser flow cytometer. ReFlex software-based quantification of energy transfer efficiency (E) directly reflects the amount of receptor interaction.
表皮生长因子（EGF）相关受体酪氨酸激酶（RTK）家族（ErbB或HER家族受体）成员在配体结合后的同源和异源相互作用是这些受体信号转导中的初始关键事件。除了各自配体的可用性外，四种ErbB受体的相对表达水平会触发受体的交叉激活，这决定了信号的多样化以及细胞的生物学反应。然而，ErbB受体及其配体的功能似乎高度复杂，其对正常细胞和肿瘤细胞的生长与增殖的影响尚未完全了解。流式细胞术荧光共振能量转移（FRET）测量有助于对受体相互作用进行定量分析。本单元使用双激光流式细胞仪，对过表达ErbB2的BT474和SK-BR-3乳腺癌细胞系中的表皮生长因子受体（EGFR，ErbB1，HER1）和ErbB2（HER2）受体相互作用进行逐细胞分析。基于ReFlex软件的能量转移效率（E）定量直接反映了受体相互作用的量。
