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Immunoassay test based on L-cysteine functionalized gold nanoparticles for direct detection of Chrondrostereum purpureum

Jaime Mejías: Instituto de Investigaciones Agropecuarias

<div><em>Chrondrostereum purpureum</em> is the causative infectious agent of silverleaf disease in different fruit trees species, involving significant economic losses in Chile. It produces wood necrosis and leaves silvering. Necrosis is evident soon after the pathogen infects wood, but foliar silvering is visible after two or more seasons. On the other hand, a non-destructive and rapid method to detect this disease is not yet available. Thus, the objective was to develop an immunoassay based on gold nanoparticles (AuNPs) for the detection of the enzyme Endopolygalacturonase (endo-PG), produced by <em>C. purpureum</em>. AuNPs with a mean diameter of 20 nm were synthesized by citrate reduction of HAuCl<sub>4</sub>, and functionalized with L-cysteine (AuNPs-Cys). Nanoparticles were characterized using TEM, UV- VIS spectroscopy, FTIR, Particle size Analyzer and Zeta potential. The optimum concentration of L-cysteine was estimated by testing a range of (0.1-4.0 µM). Antibody-coated AuNPs were prepared by mixing an AuNPs-Cys suspension and a polyclonal anti- endo-PG antibody solution. BSA was added to the mixture, incubated at room temperature and washed with PBS. The assay was able to detect endopolygalacturonase in an aqueous system. Therefore. antibody-coated AuNPs-Cys were proved to be a reliable method for the detection of the pathogen and proposed as the basis for the development of a diagnostic tool for silverleaf disease, that will reduce disease dissemination and economic losses.</div>