Encapsulation of gold nanoparticles in a chitosan nanoparticle framework via emulsion system: A key step for antigen...

Accurate diagnosis requires high precision and operational efficiency for early disease detection. Membrane-based lateral flow immunodipstick assays are practical naked-eye systems widely used for preliminary antigen (Ag) screening and typically rely on antibody (Ab)-modified gold nanoparticles (AuNPs). However, limited signal intensity remains a key challenge. Polymer-based Ab-conjugated coatings on AuNP surfaces offer a practical strategy to enhance detection performance. Chitosan (CS) is a particularly suitable polymer due to its abundant reactive functional groups, which facilitate AuNP attachment and Ab conjugation. Water-based CS (WCS), prepared via complexation with N-hydroxysuccinimide (NHS), favors the conjugation of cysteine (Cys) to form WCS-Cys nanoparticles (NPs). A comparative study between direct conjugation of WCS-Cys onto AuNP surface and the use of WCS-Cys emulsion within a toluene-in-water for in-situ reduction of HAuCl₂ demonstrates that the latter approach enables successful encapsulation of AuNPs within WCS-Cys framework, as evidenced by improved lateral flow on the immunodipstick membrane. Upon simply mixing WCS-Cys-AuNPs with Ab, efficient immobilization occurs, allowing effective lateral flow on immunodipstick without aggregation. A model assay employing Ag (rLipL32) and pathogenic leptospiral whole-cell lysates successfully validates the WCS-Cys-AuNP platform, demonstrating its potential application in Ag strip tests.