Gold Nanoparticle-Antibody Bio-Probe Analysis: Synthesis, Conjugation, Characterization and Dot Blot Assay on Paper

Gold Nanoparticle (AuNP) surface functionalized with antibodies is critical for the optimization and development of AuNP-enabled biosensing technologies. The optical properties of AuNPs and the binding specificity of antibody–antigen interactions help in the amplification of the assay signals in point-of-care technologies. Obtaining the stable AuNP–Antibody conjugate poses a major challenge as there is a chance of aggregation of the particles when the pH shifts. In this work, 37.7-nm sized spherical citrate-capped AuNPs are synthesized and are conjugated with Anti-25-OH vitamin D3 antibodies using a direct physical absorption method which requires no chemical functionalization of the AuNP or the antibody surface. The UV–vis spectroscopy and DLS outputs are used to characterize and prove the efficiency of the system. The UV–vis analysis is used to indicate the adsorption of antibodies onto the AuNP surface with the change in the absorbance peak. It was observed that the negatively charged AuNPs binded to the positive charge of the antibodies to form effective bioprobes for further detection of antigen in a short time as well as visible to the naked eye. The AuNP-Ab bioprobe’s stability and efficiency were checked with repeatability under 24 h observation.