Evolution of the PRNT: Merging tradition and innovation to set the gold standard in the era of automation.

The plaque reduction neutralization test (PRNT) is the gold standard for quantifying neutralizing antibodies owing to its high sensitivity and specificity. Widely used in seroepidemiological studies and vaccine development, PRNT is limited by labor-intensive procedures, reliance on highly skilled staff, and susceptibility to human bias during manual plaque counting. These limitations hinder reproducibility, throughput, and cross-laboratory standardization. This review summarizes recent efforts to automate PRNT workflows to address these issues. However, automation adoption is often constrained by high costs and the need for regulatory frameworks to ensure data integrity. Advancements in tailored hardware and software to diverse laboratory contexts are essential to maintaining PRNT's role in immunological assessments, bridging seroneutralizing methodologies with modern automation, and making automated methods more accessible. This review outlines the key requirements for PRNT's continued evolution.