Antibody Responses to S1-RBD and Nucleocapsid Antigens Elevate at Initial COVID-19 Infection and Remain Elevated at 12 Weeks Post Hospital Discharge

Abstract

RATIONALE:COVID-19 was first reported in Wuhan, China in December 2019 as a viral pneumonia and was found to be caused by a novel Coronavirus SARS-CoV-2. Diagnosis has included a nasopharyngeal swab examined with PCR. Immunodominant antigens have been identified for SARS-CoV-2-the Nucleocapsid and Spike-protein antigens with known genetic sequences. It is presumed that most infected with SARS-CoV-2 will develop measurable antibody titers against the Nucleocapsid and/or Spike-protein antigens as they recover from the disease. We have developed a custom Luminex assay to measure these antibodies. We hypothesize that patients undergoing treatment for COVID-19 will develop varying levels of anti-SARS-CoV-2 antibody levels after infection suggesting an acquired immunity to SARS-CoV-2.Methods:Following institutional review board approval, inpatients were recruited at the time of admission for COVID-19 care. Serum was collected at the time of hospital admission (Baseline, B, n=34) and at timepoints following hospital discharge: 2-weeks (2W, n=9), 6-weeks (6W, n=7), and 12-weeks (12W, n=4). Pooled convalescent serum from 5 PCR+ patients was used for positive control (POSCTRL) and serum collected prior to December 2019 from healthy preoperative patients (preCOVIDCtrl, n=48) were used for negative controls. IgG serum antibodies were measured via custom Luminex assay using SARSCoV-2 Spike S1 (S1-RBD) and Nucleocapsid human chimeric antibodies (HC2001 and HC2003, Genscript USA, Inc., Piscataway, NJ) to generate 5PL standard curves to convert Luminex median fluorescent intensity measurements into antibody concentrations. Concentrations for the different groups were analyzed using nonparametric comparisons for each group pair via Wilcoxon method. Statistical significance for all tests was set at p<0.05.Results:Concentrations of S1-RBD and Nucleocapsid varied widely at time of admission (B) but were significantly higher than preCOVIDCtrl (Figure 1). Elevations at later timepoints remained significantly higher for S1-RBD and Nucleocapsid than Baseline. S1-RBD concentrations were higher at 2W and 6W than Baseline. Nucleocapsid concentrations were higher at 2W than Baseline. Later timepoints for both antibodies did not differ from POS-CTRL.Conclusions:Wide variation in anti-SARS-Cov-2 antibody levels at the time of hospital admission suggest patients were in different stages of response to the infection. Antibody levels were seen to be more uniformly elevated by 2 weeks post discharge and remained elevated consistently at the 6 and 12-week time points. The negative control sera suggest no prior exposure to the SARS-CoV-2 S1-RBD antigen.