Anti sars cov 2 s assay

Serum at each time point was obtained and stored at −70 degrees Celsius if not immediately analyzed. Frozen samples were thawed for 1 h at room temperature and vortexed prior to analysis. The Roche Elecsys Anti-SARS-CoV-2 S assay (a quantitative double-antigen sandwich electro-chemiluminescent immunoassay performed on the Roche Elecsys e801 auto-analyzer) and the Snibe competitive quantitative N-Ab assay (performed on the Snibe Maglumi) have been previously described in prior studies from our laboratory [12 (link)]. We also utilized the Abbott quantitative IgG and IgM SARS-CoV-2 S-Ab assays, both of which have been previously described [14 (link),15 (link)]. The Roche total S-Ab assay reports titers in U/mL and is converted to WHO international units (BAU/mL = 0.97 × U/mL). Similarly, Abbott IgG is reported in AU/mL and is converted to WHO units (BAU/mL = 0.142 × AU/mL).

We tested serum for semiquantitative total IgG to the SARS-CoV-2 spike protein (S) receptor-binding domain with the Roche Elecsys Anti-SARS-CoV-2 S assay (anti-S IgG), qualitative detection of high-affinity antibodies to SARS-CoV-2 nucleocapsid (N) protein (anti-N IgG), and neutralizing antibodies for Wuhan D614G, Delta B.1.617.2, and Omicron B.1.1.529 strains, at LabCorp (Burlington, NC) as detailed in the Supplement. Anti-S IgG values >0.8 units per milliliter (U/mL) and neutralizing titers ≥40 inhibitory dose (ID50) (reciprocal of the sample dilution required to reduce relative luminescence units by 50%) were considered positive as previously described.³² (link) The upper limit of quantitation for anti-S IgG was 2500 U/mL. Titers below the limit of detection (LOD) were assigned a value of one-half the LOD. All time points were tested for anti-S IgG; only baseline samples were tested for anti-N IgG. Neutralizing antibodies were tested at the pre-V1, post-V2, and post-V3 or end-of-study (based on sample availability) time points in a subgroup of 60 chronologically enrolled participants.

Blood samples were collected from participants in all groups at the MEA clinic and sent to MGH to determine SARS-CoV-2 anti-S-IgG titers measured by immunoassay. Sample analysis was run at the MGH Microbiology Laboratory. The chemiluminescence enzyme immunoassay (Elecsys Anti-SARS-CoV-2 S assay) was used to analyze the antigen-specific humoral immune response, and assay results were measured on the Cobas e 601 immunoassay analyzer (Roche Diagnostics, Basel, Switzerland), with a measuring range from 0.4 U/mL to 250 U/mL (up to 25,000 U/mL with manual 1:100 dilution, and up to 50,000 U/mL with manual 1:200 dilution) [22 (link)]. All samples were processed according to the manufacturer’s instructions. Measurement results were in U/mL, with the cut-off point defined as 0.80 U/mL to differentiate samples as reactive (≥0.80 U/mL) and non-reactive (<0.80 U/mL) for anti-S-IgG [22 (link)]. The assigned U/mL were equivalent to Binding Antibody Units (BAU)/mL, as defined by the first WHO International Standard for anti-SARS-CoV-2 immunoglobulin (NIBSC code 20/136). Therefore, no conversion of units was required. In all groups, an anti-S-IgG geometric mean titer (GMT) was calculated for the whole group, among vaccinated COVID-19-naïve employees, and among those with hybrid immunity.