Clinical and Analytical Performance of an Automated Serological Test That Identifies S1/S2-Neutralizing IgG in COVID-19 Patients Semiquantitatively

Abstract

In the coronavirus (CoV) disease 2019 (COVID-19) pandemic, highly selective serological testing is essential to define exposure to severe acute respiratory syndrome CoV 2 (SARS-CoV-2). Many tests have been developed, yet with variable speeds to first results, and are of unknown quality, particularly when considering the prediction of neutralizing capacity. The LIAISON SARS-CoV-2 S1/S2 IgG assay was designed to measure antibodies against the SARS-CoV-2 native S1/S2 proteins in a standardized automated chemiluminescence assay. The clinical and analytical performances of the test were validated in an observational study using residual samples (>1,500) with a positive or negative COVID-19 diagnosis. The LIAISON SARS-CoV-2 S1/S2 IgG assay proved to be highly selective and specific and offered semiquantitative measures of serum or plasma levels of anti-S1/S2 IgG with neutralizing activity. The assay's diagnostic sensitivities were 91.3% and 95.7% at >5 or ≥15 days from diagnosis, respectively, and 100% when assessed against a neutralizing assay. The assay's specificity ranged between 97% and 98.5%. The average imprecision of the assay was a <5% coefficient of variation. Assay performance at 2 different cutoffs was evaluated to optimize predictive values. The automated LIAISON SARS-CoV-2 S1/S2 IgG assay brings efficient, sensitive, specific, and precise serological testing to the laboratory, with the capacity to test large amounts of samples per day; first results are available within 35 min, with a throughput of 170 tests/hour. The semiquantitative results provided by the test also associate with the presence of neutralizing antibodies and may provide a useful tool for the large-scale screening of convalescent-phase plasma for safe therapeutic use.

Keywords: CLIA; COVID-19; SARS-CoV-2; diagnostics; immunoassays; immunoserology; neutralization assay; neutralizing antibodies; spike.

FIG 1

Distribution of SARS-CoV-2 S1/S2 IgG levels in various patient groups. All the patient groups categorized as positive (RT-PCR positive, hospitalized, and in the ICU) are significantly different from the negative groups (RT-PCR negative, pre-COVID-19, and infected with other HCoVs) at a P of <0.0001 (***), as determined by a pairwise t test comparison with Bonferroni multiplicity adjustment. The ICU patient group is significantly different from the hospitalized patient group (###, P < 0.0001).

FIG 2

SARS-CoV-2 S1/S2 IgG measurements for 211 samples collected over the course of time from 84 patients at admission and variably thereafter up to 36 days. The upward trend was modeled using an exponential regression [ln(IgG) = A + B · exp(C · days)]. The parameter A (4.58) represents the upper limit to which the LIAISON SARS-CoV-2 S1/S2 IgG trends over time and corresponds to 98 AU/ml. A + B (1.63) is the value at time zero corresponding to 5.1 AU/ml on the original scale. The parameter C (−0.112) is the rate at which the curve moves up to the asymptote and corresponds to 1.1 AU/ml per day. The dotted horizontal line is 15 AU/ml, the positive cutoff the assay. The curve cuts this at 5 days, giving an estimate of the delay from diagnosis above which the patient is, on average, positive.

FIG 3

Distribution of the LIAISON SARS-CoV-2 S1/S2 IgG assay measurements compared to values for neutralization-positive (titers ≥ 1:40) and -negative (titers < 1:40) samples by the NT assay. Assay performance is presented specifically for neutralization using 9 and 15 AU/ml as cutoffs.

FIG 4

Relationship to and distribution of the LIAISON SARS-CoV-2 S1/S2 IgG assay levels versus NT dilutions. LIAISON SARS-CoV-2 S1/S2 IgG measurements were separated into 3 groups (<40 AU/ml, 40 to 80 AU/ml, and >80 AU/ml) and related to NT assay groups by titers: ≥1:160 (dark gray) or ≥1:80 (light gray). In the >80-AU/ml group, 33 of 38 samples have an NT assay titer of ≥1:160, while 35 of 38 samples have an NT assay titer of ≥1:80. Both titers are considered acceptable by FDA guidelines (12). The respective semiquantitative groups contain 43, 43, and 38 samples.