Validated cost-effective casein-based blocking buffers for enhanced neurocysticercosis ELISA accuracy

Accurate and affordable diagnostic tools are essential for managing neurocysticercosis, a parasitic infection of the central nervous system. In indirect ELISA, blocking buffers are crucial to prevent background noise and ensure reliable results, yet few studies have compared their performance in this specific context. This study assessed nine blocking solutions, four commercial and five prepared in-Lab, using an indirect ELISA with crude Cysticercus cellulosae antigen and a panel of 30 human serum samples (14 positive, 16 negative). By keeping all other variables constant, isolated the impact of each blocking buffer on test performance. Six formulations achieved perfect diagnostic accuracy (100 % sensitivity and specificity), while the remaining three ranged from 84.6 to 93.7 % sensitivity. ROC curve analysis confirmed high overall performance (AUC 0.957–1.000). Notably, a 3 % casein-based in-Lab blocker (B9) delivered the best results, offering strong analytical consistency, flawless diagnostic metrics, and a cost reduction of over 90 % compared to commercial alternatives. Despite the limited sample size, these findings demonstrate that well-prepared in-Lab blocking buffers, particularly those based on casein, can match or exceed commercial reagents in iELISA performance while drastically lowering costs. This approach has strong potential for expanding access to accurate NCC diagnostics in resource-limited settings.