Dot Blot Validation of Eluted Schistosoma Immunogenic Proteins for Rapid Schistosomiasis Diagnosis

Schistosomiasis remains a major public health concern in endemic regions where rapid and accurate diagnosis is vital for effective control. Persistent diagnostic challenges arise as a result of crude parasite antigen-based assays, which often provoke non-specific immune responses and compromise accuracy. This study validated two immunoreactive proteins eluted from Schistosoma haematobium soluble egg antigen (SEA) and Schistosoma mansoni worm antigen proteins (WAPs) as improved diagnostic tools for field deployment using dot blot assays.

SDS–polyacrylamide gel electrophoresis and Western blot assays identified immunoreactive protein bands from SEA and WAP, which were then evaluated using ELISA on 40 repository samples (20 positive and 20 negative). The bulk of the validation work employed dot blot assays, which is the primary qualitative method used to demonstrate clear immunogenicity of each antigen against patient urine. Statistical analyses, including tests of mean optical density, area under ROC curve (AUC), sensitivity, and specificity, were performed using GraphPad Prism 8.0.1 and MedCalc webserver.

Two highly immunoreactive proteins were successfully eluted: 72 kDa from Sh SEA and 95 kDa from Sm WAP, with diagnostic cut-offs of 0.3383 and 0.1102, respectively. The 72 kDa protein demonstrated excellent performance with 95% sensitivity and 90% specificity, while the 95 kDa protein showed 90% sensitivity and 95% specificity. Both exhibited excellent diagnostic accuracy of 72 kDa (AUC = 0.96; 95% CI: 0.90–1.00) and 95 kDa (AUC = 0.96; 95% CI: 0.91–1.00). Dot blot validation clearly revealed the immunogenic potential of both proteins, visually differentiating positive from negative samples. ELISA results showed significant differences in mean optical density between positive (0.75±0.27), (0.35±0.19) and negative (0.23±0.15), (0.07±0.02) samples (P <0.0001).

The 72 kDa and 95 kDa eluted proteins demonstrated strong immunogenicity and diagnostic precision, addressing the limitations of crude antigen-based assays and offering more specific and reliable tools for schistosomiasis detection in endemic regions.