title: Quantification of epidermal growth factor receptor expression level and binding kinetics on cell surfaces by surface plasmon resonance imaging.
abstract: Epidermal growth factor receptor (EGFR, also known as ErbB-1 or HER-1) is a membrane bound protein that has been associated with a variety of solid tumors and the control of cell survival, proliferation, and metabolism. Quantification of the EGFR expression level in cell membranes and the interaction kinetics with drugs are thus important for cancer diagnosis and treatment. Here we report mapping of the distribution and interaction kinetics of EGFR in their native environment with the surface plasmon resonance imaging (SPRi) technique. The monoclonal anti-EGFR antibody was used as a model drug in this study. The binding of the antibody to EGFR overexpressed A431 cells was monitored in real time, which was found to follow the first-order kinetics with an association rate constant (ka) and dissociation rate constant (kd) of (2.7 ± 0.6) × 10(5) M(-1) s(-1) and (1.4 ± 0.5) × 10(-4) s(-1), respectively. The dissociation constant (KD) was determined to be 0.53 ± 0.26 nM with up to seven-fold variation among different individual A431 cells. In addition, the averaged A431 cell surface EGFR density was found to be 636/μm(2) with an estimation of 5 × 10(5) EGFR per cell. Additional measurement also revealed that different EGFR positive cell lines (A431, HeLa, and A549) show receptor density dependent anti-EGFR binding kinetics. The results demonstrate that SPRi is a valuable tool for direct quantification of membrane protein expression level and ligand binding kinetics at single cell resolution. Our findings show that the local environment affects the drug-receptor interactions, and in situ measurement of membrane protein binding kinetics is important.
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.|School of Electrical Computer and Energy Engineering, Arizona State University , Tempe, Arizona 85287, United States.
authors.forename: Fenni
authors.initials: F
authors.lastname: Zhang
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.
authors.forename: Shaopeng
authors.initials: S
authors.lastname: Wang
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.|College of Chemistry and Chemical Engineering, Chongqing University , Chongqing 400044, China.
authors.forename: Linliang
authors.initials: L
authors.lastname: Yin
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.|School of Electrical Computer and Energy Engineering, Arizona State University , Tempe, Arizona 85287, United States.
authors.forename: Yunze
authors.initials: Y
authors.lastname: Yang
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.|School of Electrical Computer and Energy Engineering, Arizona State University , Tempe, Arizona 85287, United States.
authors.forename: Yan
authors.initials: Y
authors.lastname: Guan
authors.affiliation: State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University , Nanjing 210093, China.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: Amgen Inc. , Thousand Oaks, California 91320, United States.
authors.forename: Han
authors.initials: H
authors.lastname: Xu
authors.affiliation: Center for Bioelectronics and Biosensors, Biodesign Institute, Arizona State University , Tempe, Arizona 85287, United States.|School of Electrical Computer and Energy Engineering, Arizona State University , Tempe, Arizona 85287, United States.|State Key Laboratory of Analytical Chemistry for Life Science, School of Chemistry and Chemical Engineering, Nanjing University , Nanjing 210093, China.
authors.forename: Nongjian
authors.initials: N
authors.lastname: Tao
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology*; D002460:Cell Line; D004867:Equipment Design; D066246:ErbB Receptors / Q000032:analysis* / Q000276:immunology*; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D020349:Surface Plasmon Resonance / Q000295:instrumentation*
chemical_list: D000911:Antibodies, Monoclonal; C512478:EGFR protein, human; D066246:ErbB Receptors