Development of Au-Pt Nanoflowers as Nanoenzyme for Highly Sensitive Immunoassay: Application to the Detection of Melioidosis Biomarker
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Authors
Kharal, Gita
Issue Date
2024
Type
Thesis
Language
Keywords
Au-Pt Nanoflowers , Bio-conjugation , Enzyme Kinetics And Optimization , Immunoassay , Melioidosis Biomarker , Nanoenzyme
Alternative Title
Abstract
Immuno-sensing techniques, which are well-known for their high efficiency and selectivity, have become widely used for detecting biomarkers in clinical and research settings. Nanoparticles with enzyme-mimic properties, known as nanozymes, have garnered significant attention for their potential in revolutionizing immuno-sensing due to their tunable catalytic activity, well-developed synthesis techniques, and stability across a wide range of pH, temperature, and inhibitor concentration. Among these, Au-Pt nanostructures exhibit exceptional peroxidase-mimic catalytic activity, making them promising candidates for enhancing immunoassay techniques such as ELISA and LFIA. In this work, different nanoenzymes and their use in immunoassay application were reviewed. In this study, we present a core shell gold-platinum nanoflowers (Au-Pt NFs) as signal labels in immunoassays for sensitive detection of capsular polysaccharides (CPS) expressed on the cell envelopes of Burkholderia pseudomallei causing Melioidosis, an infectious disease of global health concern. Utilizing the inherent peroxidase-like activity of Au-Pt NFs, we developed Nano-Enzyme Linked Immunosorbent Assay (N-ELISA) and Lateral Flow Immunoassay (LFIA) techniques for on-site and sensitive CPS detection. Au-Pt NFs were synthesized by growing Pt nanoflowers on gold nanoparticles (AuNPs) surfaces, ensuring dual functionality as signal labels. Through the use of a developed nanoenzyme, lateral flow assays for CPS were made possible using the Au-Pt NFs conjugated with anti-CPS antibodies. By only using the Au-Pt NFs without sophisticated instruments, it was possible to visually detect the test and control lines. Through colorimetric enhancement, our Au-Pt NF-based LFIA demonstrated superior sensitivity, detecting CPS at a minimum concentration of ~70 pg/mL, surpassing previous AuNP-based LFIA methods. Clinical diagnostic utility was validated by successfully detecting CPS spiked in human serum samples (~100 pg/mL).The utilization of Au-Pt NFs as signal labels in immunoassays offers a promising avenue for enhancing diagnostic capabilities, particularly in infectious diseases like Melioidosis. By overcoming the limitations of conventional enzyme-based assays, Au-Pt NFs pave the way for rapid and accurate point-of-care diagnostics.