Using optical waveguide-based particle plasmon resonance biosensor for quantitative measurement of biomolecular interaction kinetics

• Main Authors: Wu, Chao-Ching, 吳昭慶
• Other Authors: Chau, Lai-Kwan
• Format: Others
• Language: zh-TW
• Published: 2011
• Online Access: http://ndltd.ncl.edu.tw/handle/05425759914430516160

碩士 === 國立中正大學 === 化學暨生物化學研究所 === 99 === Measuring the kinetic constants of antigen-antibody interactions becomes critical in characterizing specific affinity and, hence, exploring the feasibility of using such interactions in clinical diagnosis. In this study, we used the optical waveguide-based particle plasmon resonance biosensor for quantitative measurement of biomolecular interaction kinetics. For the first part, two configurations of biosensing chips were used. Chip1 has a one-dimensional surface grating structure with a period of 833 nm by stamping with UV glue on a glass slide and gold nanoparticles are modified on the opposite side of the glass slide. Chip2 has a grating structure with a period of 833 nm by stamping on a photopolymerizable sol-gel layer which is coated on a glass slide and gold nanoparticles are modified on the sol-gel layer. The planar waveguides for chip1 and chip2 are the glass slide and the photopolymerized sol-gel layer, respectively. Detection of the optical resonant reflective position from waveguide surface enables high sensitivity label-free biosensing. The light source in this setup was a diode-pumped laser (532nm). The sensor resolution achieved by the gold nanoparticles-modified biosensing chips was 2.8×10-5 RIU for chip1 and 1.1×10-4 RIU for chip2. The detection limit for anti-DNP antibody was determined to be 1.4×10-11 M for chip1 and 5.4×10-10 M for chip2. Unfortunately, even we could observe the signal-time sensorgram, we could not measure the kinetic constants because of the low S/N ratio achieved by these optical setups. For the second part, a simple and label-free biosensing method has been developed for the determination of anti-ovalbumin antibody (anti-OVA) and anti-Mouse IgG antibody and monitoring their biomolecular interactions by using the fiber-optic particle plasmon resonance (FOPPR) biosensor. The FOPPR sensor is based on gold nanoparticles-modified optical fiber where the gold nanoparticle surface has been modified by a mixed self-assembled monolayer for conjugation of receptor (ovalbumin or mouse IgG) to detect the corresponding analyte (anti-OVA or anti-mouse IgG). During the detection process, an analyte reacts with a receptor immobilized on the optical fiber and their interaction can be monitored in real-time. The detection limits for anti-OVA antibody and anti-mouse IgG were determined to be 2.4×10-9 M and 4.3×10-9 M, respectively. In addition, by assuming a Langmuir type of adsorption isotherm, the quantitative measurement of binding kinetics, including the association and dissociation rate constants, yields a ka of 5.5×103 M-1s-1 and a kd of 3.5×10-4 s-1 for OVA/anti-OVA, and a ka of 1.9×106 M-1s-1 and a kd of 1.7×10-2 s-1 for mouse IgG/anti-mouse IgG. We demonstrate that the FOPPR biosensor can study real-time biomolecular interactions.