Immunoglobulin Binding Analysis by Utilizing a Surface Plasmon Resonance Phase-Measurement System

• Main Authors: Shiuan Lin, 林萱
• Other Authors: Yi-Chun Chen
• Format: Others
• Language: zh-TW
• Published: 2012
• Online Access: http://ndltd.ncl.edu.tw/handle/64704117083856689559

碩士 === 國立中央大學 === 照明與顯示科技研究所 === 100 === A dynamic surface-plasmon-resonance (SPR) sensor was developed based on interferometric phase measurements. Temporal intensity signals were acquired by photodetectors and processed by the Schwider-Hariharan five-step algorithm to obtain the phases. The achieved phase-detection stability was 1.2214 degrees in a 3600-second period and the system sensitivity was 1.1465×10^4 degree/RIU (refractive index unit). The corresponding system resolution was 1.0653×10^-4 RIU. Salt-water mixture measurements were performed to characterize the relation between the measured phase difference and the refractive-index variation of the specimen. Several known concentrations of salt-water mixtures were flowed into the SPR system to measure the phases. A refractometer was utilized to measure the refractive indices. The results showed a linear correspondence between the phase difference and the refractive index. The measurement of an unknown salt-water mixture showed a relative refractive-index error of about 0.023 % between the refractometer measurement and the linear interpolation result from the SPR system. For the antibody-antigen binding experiments, the phase differences caused by the injection of anti-IgG with concentrations of 25 μg/ml, 50 μg/ml, 100 μg/ml and 200 μg/ml were measured to be approximately 33.1399°, 41.0506°, 58.8533°, and 78.7476°, respectively. From the relation between the time and phase difference, the association rate constant ka and the dissociation rate constant kd were calculated to be 4.1139×10^4 M^-1s^-1 and 2.8768×10^-4 s^-1, respectively. By utilizing the relation between the SPR signals and the concentrations of the analyte from salt-water mixture measurements and antibody-antigen binding experiments, the concentration of unknown solutions can be estimated by this SPR system.