A combined strategy to detect plasma samples reliably with high anti-SARS-CoV-2 neutralizing antibody titers in routine laboratories

Abstract

The determination of anti-SARS-CoV-2 neutralizing antibodies (NAbs) is of interest in many respects. High NAb titers, for example, are the most important criterion regarding the effectiveness of convalescent plasma therapy. However, common cell culture-based NAb assays are time-consuming and feasible only in special laboratories. Our data reveal the suitability of a novel ELISA-based surrogate virus neutralization test (sVNT) to easily measure the inhibition-capability of NAbs in the plasma of COVID-19 convalescents. We propose a combined strategy to detect plasma samples with high NAb titers (≥ 1:160) reliably and to, simultaneously, reduce the risk of erroneously identifying low-titer specimens. For this approach, results of the sVNT assay are compared to and combined with those acquired from the Euroimmun anti-SARS-CoV-2 IgG assay. Both assays are appropriate for high-throughput screening in standard BSL-2 laboratories. Our measurements further show a long-lasting humoral immunity of at least 11 months after symptom onset.

Evaluation of commercial Anti-SARS-CoV-2 neutralizing antibody assays in seropositive subjects

“…It can be postulated that part of the antibodies detected by these ELISA are able to interfere with the interaction between ACE receptor and the viral RBD but not to prevent cell entry of the virus; this may be related to the affinity/avidity of antibodies that is reported to be low after primary infection or first vaccine dose ( 11 , 12 ), and is likely to be even more the case for the population included herein who were sampled 6 months after infection. Despite this low specificity, these assays correlated with the live virus neutralization assay as also found in other studies ( 5 , 6 , 13 , 14 , 15 ). The low specificity could also be attributed to a lack of sensitivity of live VNT but it is rather unlikely that decreasing the threshold below 20 would be clinically relevant, such low titer having little chance to be protective in vivo ( 16 ).…”

“…The study of Von Rein et al ( 22 ) suggested that the correlation between sVNT and VNT was greater at higher level of neutralization titer and they concluded that sVNT are only useful when inhibition was above 50%, which is more consistent with our data. Most of the previous studies used the cPASS assay from GeneSript,showing the correlation of competitive immunoassay to live VNT, with good sensitivity and specificity compared to VNT ( 5 , 6 , 9 , 14 , 15 , 19 , 20 , 22 , 23 ). Only a few studies reported results with the TECO ( 15 , 20 ) or YHLO assays ( 7 , 8 , 21 ).…”

“…Other studies have compared VNT and sVNT with assays detecting IgG binding to RBD or S proteins and observed a correlation between them (6,8,9). Fisher et al ( 6 ) found that the correlation between sVNT and antibody binding assay is better for samples with high than low PRNT. Others studies have shown that the correlation between VNT and antibody binding assays was lower than between sVNT and antibody binding assays ( 9 ).…”

“…These competitive immunoassays, which can be conducted using qualitative immunochromatographic cassettes or quantitative automated or manual enzyme-linked immunosorbent assay (ELISA) platforms, allow rapid and easy processing of large numbers of samples in conventional serological laboratories. However, the performance of these newly developed commercial sVNT assays by comparison to classical serological assays detecting anti-RBD IgG and/or to the reference plaque reduction neutralization test 50% (PRNT 50 ) performed with live virus has been poorly evaluated up to now ( 6 , 7 , 8 , 9 ). Moreover, previous studies have evaluated the specificity using seronegative and/or prepandemic serum which do not inform if commercial sVNT can differentiate serum with or without neutralizing antibody in seropositive samples.…”

Evaluation of Commercial Anti-SARS-CoV-2 Neutralizing Antibody Assays in seropositive subjects

“…It can be postulated that part of the antibodies detected by these ELISA are able to interfere with the interaction between ACE receptor and the viral RBD but not to prevent cell entry of the virus; this may be related to the affinity/avidity of antibodies that is reported to be low after primary infection or first vaccine dose (11,12), and is likely to be even more the case for the population included herein who were sampled 6 months after infection. Despite this low specificity, these assays correlated with the live virus neutralization assay as also found in other studies (5,6,13–15). The low specificity could also be attributed to a lack of sensitivity of live VNT but it is rather unlikely that decreasing the threshold below 20 would be clinically relevant, such low titer having little chance to be protective in vivo (16).…”