A proof-of-concept colloidal gold nanoparticle-based immunochromatographic dip strip for rapid detection of the Staphylococci group in mastitic dairy milk

Mastitis caused by staphylococci represents a major challenge to the health of dairy cattle and the quality of milk worldwide, underscoring the need for rapid and reliable diagnostic tools. This study describes the development of a colloidal gold nanoparticle-based immunochromatographic dip strip for the detection of staphylococcal bacteria in dairy milk. Monoclonal antibodies were conjugated to 40-nm colloidal gold nanoparticles at pH 8.2, with various membrane types and buffer compositions evaluated to ensure optimal performance. At a concentration of 10 9 CFU/mL, the strip successfully identified Staphylococcus aureus and several Non-Aureus Staphylococci (NAS) species, producing a distinct red signal at the test dot, while no cross-reactivity was observed with non-staphylococcal pathogens. The assay achieved a limit of detection (LOD) of 10 8 CFU/mL in ultra high temperature (UHT) milk, with results available within 20 minutes. Stability testing demonstrated that the strips remained functional for up to 30 days when stored at 25 °C in aluminum foil packaging. Although the current LOD is primarily suitable for samples with a high bacterial load, this study provides critical proof of concept for a rapid, genus-specific screening platform. The developed strip offers a practical, on-site solution for monitoring high-burden staphylococcal infections, serving as a valuable tool for immediate herd management and mastitis control.