Brucellosis is a zoonotic disease endemic in developing countries and caused by gram-negative bacteria of genus Brucella infecting both livestock and humans. Brucella melitensis and Brucella abortus are important species representing largest biotypes world-wide. Therefore, modified detection strategies and advancement in potential analytical tools are required to monitor its rapid prevalence. In this study, we aim to modify gold-transducer of Surface Plasmon Resonance biosensor with combination of metal oxide nanomaterials and chemical probe to detect recombinant outer membrane ‘rOmp28’ protein antigen of Brucella melitensis 16M in concentration-dependent surface interactions. We synthesized Zinc (ZnONPs) and Gold (AuNPs) nanoparticles using standard ‘Hydrothermal and Turkevich Methods’ and their crystalline structure, chemical property and morphology were analysed using UV-Visible Spectrophotometry, FT-IR, Powder-XRD, SEM-EDX and TEM-SAED. For immobilizing specific rOmp28 derived IgG-pAbs on modified Au-transducer, rOmp28 protein was expressed and purified using Ni-NTA gel affinity chromatography for producing pAbs in BALB/c mice. Three modifications of Au-transducer with 4-MBA, ZnONPs/AuNPs and ZnONPs/AuNPs/4-MBA were subjected for immobilization and SPR biosensing was performed with rOmp28 Ag at detection range of 0.1 μg mL-1 to 0.01 fg mL-1. Limit of detection observed with ZnONPs/AuNPs/4-MBA combination was 0.1 fg mL-1 by relative increase in SPR response angle at 0.1 μg mL-1 in the order 83.7° < 98.9° < 179.2° for 4-MBA < ZnONPs/AuNPs < ZnONPs/AuNPs/4-MBA respectively. In conclusion, metal oxide nanomaterials in combination with biosensor are suitable in sensitive and specific interaction of antigen displaying lowest LODs and enhanced biosensor response for on-field real-time Brucella detection in both clinical and non-clinical disease scenario.
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