Rapid Detection of Sheep-Specific <i>Clostridium perfringens</i> IgG Using Silver Nanoparticle-Boosted Dot-Blot Assays

Dot-blot immunoassay is a quick, simple, and low-cost technique for identifying target analytes in food, clinical diagnostics, and the environment with a visual inspection. For this purpose, silver nanoparticles (AgNPs) were synthesized using a sodium borohydride compound. Synthesized AgNPs were characterized using UV-Vs spectroscopy, scanning electron microscope (SEM), X-ray diffraction (XRD), dynamic light scattering (DLS), and zeta potential (ZP) tests. After that, SNPs were conjugated with anti-CLD N4-C. perfringens IgG. At the same time, the positive and negative C. perfringens samples have been determined using ELISA. Furthermore, PCR test was used for verification of C. perfringens. Consequently, dot-blot assay was performed on nitrocellulose membranes using ultrasonicated C. perfringens antigen. On specific ELISA (golden-test) examination, 39 of the 100 samples were positive for C. perfringens, and 61/100 were negative. On the nitrocellulose membrane the reaction was occur during 1–2 min. The existence of a blue color blot verified the positive reaction. 5/61 dot-blot tests were false negative, indicating 91.80% specificity, while 7/39 were false positive, indicating 82.5% sensitivity. Dot-blot sensitivity and specificity indicated 88% accuracy. The ELISA-dot blot concordance was k = 81.2% (k value = 0.62), which is a significant agreement. Furthermore, after tenfold serial dilution of 100 μg/ml, 10 μg/ml, 1 μg/ml, and 0.1 μg/ml, the sensitivity of positive dot-blot samples was 100%, 93%, and 62%, respectively. The AgNP-dot-blot assay showed high sensitivity, specificity, and accuracy for detecting C. perfringens, making it an easy, rapid, economical, and accurate diagnostic tool for identifying enterotoxaemia in sheep.