Site-Specific Immobilization of Antibody on Gold Nanoparticle Surface for One-Step Diagnosis of Prostate Specific Antigen with Dynamic Light Scattering

Small dimensions of gold nanoparticles (AuNPs) necessitate the oriented immobilization of antibodies to conserve antibodies’ antigen binding activity for proper function. In this study, we used four different methods to immobilize antibodies on AuNP surface; i) UV-NBS, ii) EDC-NHS, iii) disulfide reduction for thiol conjugation chemistry, and iv) physical adsorption; and compared the efficiency of these methods in terms of antigen binding activity, detection sensitivity and selectivity. Two different clones of anti-PSA monoclonal antibodies, B731M and B728M, specific to different epitopes on Prostate Specific Antigen (PSA) were chosen to functionalize on 5 nm AuNPs. Once the mixture of two different bivalently active antibodies on AuNPs recognize PSA, they form large clusters depending on the concentration of PSA in a sample; size of those clusters can be easily measured by dynamic light scattering (DLS) allowing selective detection of the antigen. Our results demonstrate that the limit of detection (LOD) for PSA utilizing antibody functionalized AuNPs via the UV-NBS method was significantly lower than the other immobilization methods, with the LOD of 0.1 nM PSA. The UV-NBS method yielded AuNP surfaces with significantly enhanced antigen recognition capabilities and improved antigen detection sensitivity with high level of selectivity when compared to other commonly used AuNP functionalization methods including: EDC-NHS, disulfide reduction for thiol conjugation chemistry, and physical adsorption. Consequently, this study demonstrates a one-step PSA detection method that yields highly sensitive, selective and rapid results.