NIH Researchers outline the development of an ELISA-based serology assay protocol that measures antibodies against coronavirus proteins to better predict the prevalence and understand the spread of COVID-19. The protocol standardizes thresholds of IgG, IgM, and IgA antibodies in multiple sample types with high specificity and sensitivity. The researchers used purified antigens to compare to currently available antigens to determine which combination is the most sensitive to SARS-CoV-2. Through careful statistical evaluations, the researchers determined that using a combination of the McLellan/VRC spike construct and the Ragon RBD in the assay gave the most sensitive results. While there is a manual protocol, the researchers also developed a semi-automated protocol that increases the speed of processing and standardization to maintain consistency and increase throughput. Both methods resulted in a 100% sensitivity and specificity at three standard deviations above negative control values. This protocol utilizes common reagents and instruments such that this protocol can be easily adapted to other labs. However, validation steps must be done to account for variances in equipment.