04670nas a2200205 4500008004100000245011000041210006900151260000800220300001400228490000700242520391400249100002104163700002204184700002404206700002304230700003404253700002704287700002204314856012804336 2019 eng d00a Development of a nano-gold immunodiagnostic assay for rapid on-site detection of invasive aspergillosis 0 aDevelopment of a nanogold immunodiagnostic assay for rapid onsit cSEP a1341-13520 v683 a

Introduction. Timely detection of invasive aspergillosis (IA) caused by fungal pathogens, i.e. Aspergillus fumigatus and Aspergillus flavus, in immunocompromised patients is crucial in preventing high mortality.

Aim. To develop a simple immunoassay for the detection of galactomannan (GM), an IA biomarker.

Methodology. GM from A. fumigatus and A. flavus clinical strains was purified and characterized by X-ray diffraction, IR spectroscopy and C-13/H-1 nuclear magnetic resonance (NMR) for polyclonal antibody (pAb) production in rabbits. An enzyme-linked immunosorbent assay (ELISA) was standardized using concanavalin A to capture Aspergillus GM and pAbs to detect it. Gold nanoparticles (AuNPs) were synthesized and conjugated to pAbs for the development of a dot-blot immunoassay. The developed dot-blot was evaluated with 109 clinical serum and bronchoalveolar lavage samples.

Results. Spectroscopy studies characterized the D-galactofuranosyl groups of GM responsible for the immune response and generation of pAbs. The ELISA employing pAbs showed a sensitivity of 1 ng ml(-1) for Aspergillus GM. Furthermore, a sensitive, visual, rapid dot-blot assay developed by the conjugation of pAbs to AuNPs (similar to 24 +/- 5 nm size, -36 +/- 2 mV zeta potential) had a detection limit of 1 pg ml(-1) in serum. The pAbs interacted with Aspergillus spp. but did not cross-react with other fungal pathogen genera such as Penicillium and Candida. Evaluation of the dot-blot with 109 clinical samples showed high sensitivity (80%) and specificity (93.2 %), with an overall assay accuracy of 89%.

Conclusion. The developed nano-gold immunodiagnostic assay has immense potential for practical use in rapid, specific and sensitive on-site diagnosis of IA, even under resource-limited settings.

1 aRaval, Komal, M.1 aGhormade, Vandana1 aRajamohanan, P., R.1 aChoudhary, Hansraj1 aRudramurthy, Shivaprakash, M.1 aChakrabarti, Arunaloke1 aPaknikar, Kishore uhttps://library.ncl.res.in/content/development-nano-gold-immunodiagnostic-assay-rapid-site-detection-invasive-aspergillosis