D'Abronzo Leandro S, Pan Chong-Xian, Ghosh Paramita M
VA Northern California Health Care System, University of California at Davis, Sacramento, CA, USA.
Department of Urology, University of California at Davis, Sacramento, CA, USA.
Methods Mol Biol. 2018;1655:319-334. doi: 10.1007/978-1-4939-7234-0_22.
The epidermal growth factor receptor (EGFR) family of receptor tyrosine kinases (RTK) consists of four members: EGFR1/ErbB1/HER1, ErbB2/HER2, ErbB3/HER3, and HER4/ErbB4. Signaling through these receptors regulates many key cellular activities, such as cell division, migration, adhesion, differentiation, and apoptosis. The ErbB family has been shown to be overexpressed in different types of cancers and is a target of several inhibitors already in clinical trials. ErbB3 lacks a functional tyrosine kinase domain and therefore has not been as extensively studied as the other members of this family, but its importance in activating downstream pathways, such as the PI3K/Akt pathway, makes this RTK a worthy investigation target, especially in urothelial carcinoma where the PI3K/Akt pathway is vital for progression. In recent times, ErbB3 overexpression has been linked to drug resistance and progression of various diseases, especially cancer. ErbB3 levels in the serum were shown in many cases to be reflective of its role in disease progression, and therefore detection of serum ErbB3 levels during treatment may be of importance.Here we describe two methods for detecting ErbB3 protein in serum from patients who have undergone a clinical trial, utilizing two well-established methods in molecular biology-western blotting and ELISA, focusing on sample preparation and troubleshooting.
受体酪氨酸激酶（RTK）的表皮生长因子受体（EGFR）家族由四个成员组成：EGFR1/ErbB1/HER1、ErbB2/HER2、ErbB3/HER3和HER4/ErbB4。通过这些受体发出的信号调节许多关键的细胞活动，如细胞分裂、迁移、黏附、分化和凋亡。已证明ErbB家族在不同类型的癌症中过度表达，并且是几种已进入临床试验的抑制剂的靶点。ErbB3缺乏功能性酪氨酸激酶结构域，因此没有像该家族的其他成员那样得到广泛研究，但其在激活下游通路（如PI3K/Akt通路）中的重要性使这种RTK成为一个值得研究的靶点，特别是在PI3K/Akt通路对进展至关重要的尿路上皮癌中。近年来，ErbB3的过度表达与各种疾病尤其是癌症的耐药性和进展有关。在许多情况下，血清中的ErbB3水平反映了其在疾病进展中的作用，因此在治疗期间检测血清ErbB3水平可能很重要。在这里，我们描述了两种用于检测参与临床试验患者血清中ErbB3蛋白的方法，利用分子生物学中两种成熟的方法——蛋白质印迹法和酶联免疫吸附测定法，重点关注样品制备和故障排除。
