The role of the interfacial design of nanoparticles in the performance of optical single molecule biosensor

How to design a biosensor with optimal performance with regards to sensitivity, specificity, response time and the ability to be multiplexed is a key issue to meet the demands of diagnostics for health cares. As modern biosensors are usually constructed with bioreceptors immobilized on nanoparticles, the surface of nanoparticle has a significant impact on sensing performance. This thesis aims at investigating how to design the surface of nanoparticle to achieve better sensing performance of single molecule biosensor. Two different biosensing systems were built using DNA functionalized gold nanoparticle (AuNPs), including fluorescent biosensor and localized surface plasmon resonance (LSPR) biosensor using dark-field microscopy. To better understand how the coverage of DNA on surface of AuNPs affects sensing performance, the coverage of DNA on AuNPs surface was adjusted and effect of coverage of DNA on the sensing performance was investigated. Firstly, the impact of coverage of aptamer on the surface of AuNPs on sensing performance was investigated using fluorescence spectroscopy. The number of anti-interferon gamma (IFN-) aptamers was adjusted from an average of 9.6 to 258 per nanoparticle. The sensing strategy was based on the conformational change of anti-IFN- aptamer and fluorescence quenching property of AuNPs. The binding isotherm and binding kinetics of the interaction between AuNPs-aptamer conjugate and IFN- as a function of coverage of aptamer were investigated. It was found that AuNPs-aptamer conjugate with the highest coverage of aptamer was the most favorable in biosensors considering the limit of detection, sensitivity, and response time of detection. Secondly, a LSPR biosensor utilizing AuNPs-DNA conjugates was built to detect SARS-CoV-2 specific RNA. Two AuNPs-DNA conjugates with different size of AuNPs were adopted to form a dimer consisting with a 80 nm gold nanoparticle and a 40 nm gold nanoparticle. The binding of target RNA with dimer resulted in colour change of dimer under dark-field microscope. A calibration curve between response and different concentration of target RNA was obtained with a determined limit of detection of 2.6 fM. Lastly, the number of DNA on the satellite nanoparticle was adjusted by different diluents to optimize the sensing performance of LSPR biosensor. By reducing the amount of DNA on 40 nm gold nanoparticle, the response for the same concentration of target RNA was improved and lower limit of detection was achieved. This thesis provides insights into how to design the surface of nanoparticle and the impact of interfacial design of nanoparticle on the sensing performance of biosensor, which advances the development of single molecule biosensor from the perspective of interface of nanoparticle