Evaluation of commercially available fully automated and ELISA-based assays for detecting anti-SARS-CoV-2 neutralizing antibodies

Abstract

Rapid and accurate measurement of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV2)-specific neutralizing antibodies (nAbs) is paramount for monitoring immunity in infected and vaccinated subjects. The current gold standard relies on pseudovirus neutralization tests which require sophisticated skills and facilities. Alternatively, recent competitive immunoassays measuring anti-SARS-CoV-2 nAbs are proposed as a quick and commercially available surrogate virus neutralization test (sVNT). Here, we report the performance evaluation of three sVNTs, including two ELISA-based assays and an automated bead-based immunoassay for detecting nAbs against SARS-CoV-2. The performance of three sVNTs, including GenScript cPass, Dynamiker, and Mindray NTAb was assessed in samples collected from SARS-CoV-2 infected patients (n = 160), COVID-19 vaccinated individuals (n = 163), and pre-pandemic controls (n = 70). Samples were collected from infected patients and vaccinated individuals 2–24 weeks after symptoms onset or second dose administration. Correlation analysis with pseudovirus neutralization test (pVNT) and immunoassays detecting anti-SARS-CoV-2 binding antibodies was performed. Receiver operating characteristic (ROC) curve analysis was generated to assess the optimal threshold for detecting nAbs by each assay. All three sVNTs showed an excellent performance in terms of specificity (100%) and sensitivity (100%, 97.0%, and 97.1% for GenScript, Dynamiker, and Mindray, respectively) in samples collected from vaccinated subjects. GenScript demonstrated the strongest correlation with pVNT (r = 0.743, R2 = 0.552), followed by Mindray (r = 0.718, R2 = 0.515) and Dynamiker (r = 0.608, R2 = 0.369). Correlation with anti-SARS-CoV-2 binding antibodies was variable, but the strongest correlations were observed between anti-RBD IgG antibodies and Mindray (r = 0.952, R2 = 0.907). ROC curve analyses demonstrated excellent performance for all three sVNT assays in both groups, with an AUC ranging between 0.99 and 1.0 (p < 0.0001). Also, it was shown that the manufacturer's recommended cutoff values could be modified based on the tested cohort without significantly affecting the sVNT performance. The sVNT provides a rapid, low-cost, and scalable alternative to conventional neutralization assays for measuring and expanding nAbs testing across various research and clinical settings. Also, it could aid in evaluating actual protective immunity at the population level and assessing vaccine effectiveness to lay a foundation for boosters' requirements.

Novel neutralizing mouse-human chimeric monoclonal antibodies against the SARS-CoV-2 receptor binding domain

“…Our results and those reported by other investigators indicate lower sensitivity of the sVNT as compared to the pVNT and cVNT methods [47, 48]. Despite these differences the results reported in vaccinated and convalescent patients show a positive correlation between these three viral neutralization tests [49, 50]. The IC50 values reported for the commercial FDA approved therapeutic mAbs are within the range of 0.001- to 0.058 µg ml −1 [44].…”

Importance, Applications and Features of Assays Measuring SARS-CoV-2 Neutralizing Antibodies

“…Based on the knowledge that the SARS-CoV-2 virus infects the host cells by docking via a portion of the viral S protein termed RBD to its cognate receptor ACE2 on the host cells [ 11 ], molecular interaction assays were developed which allowed us to investigate if antibodies from convalescent subjects can inhibit the RBD–ACE2 interaction and thus have virus-neutralizing capacity [ 12 , 13 , 14 ]. In fact, results from the molecular interaction assays correlated very well with results obtained by classical virus-neutralization tests [ 15 , 16 , 17 , 18 ].…”

Development of an IgY-Based Treatment to Control Bovine Coronavirus Diarrhea in Dairy Calves

“…Additionally, to confirm a correlation between VN titers and ELISA BCoV-specific IgY titers we performed a regression analysis with log−10 transformed titers resulting in an R 2 value of 0.92 for a linear regression analysis and 0.97 for the quadratic regression analysis ( Supplementary File S1 ). These results are very optimistic compared to similar studies performed with a correlation analysis between GenScript SARS-CoV-2 ELISA Ab titers and pseudo-VN titers with an R 2 value of 0.552 [ 45 ]. This is the first BCoV-specific IgY ELISA assay to be statistically validated.…”