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Figure 1 . SARS-CoV-2 S protein including the ectodomain (residues 19-1207) without native signal peptide. The furin cleavage site was removed (GSAS mutations) and residues at positions 986 (K) and 987(V) were mutated to proline. The C-terminal included a 4 fibritin trimerization domain, a TEV protease cleavage site, and a histidine tag.
Figure 2 . Rachiplusia nu larvae. a. A batch of 500 fifth-instar larvae. b. Injection of the recombinant baculovirus near the third prolegs.
Figure 3. Purification of S protein by IMAC. a. SDS-PAGE with Coomassie blue staining. b. Western blot with DAB staining. Lanes: MK. molecular weight marker. 1. Larval extract (input). 2. Passthrough. 3. Wash fraction with 80 mM imidazole. 4. Elution fraction with 500 mM imidazole.
Figure 4. Size exclusion analysis of S protein purified by IMAC. a) Chromatogram showing the elution profile of S protein in IMAC. The void volume (Blue dextran) of the column is ~8.8 ml, and the monomeric (~67 kDa) and dimeric (~140 kDa) forms of Bovine Serum Albumin (BSA) eluting at ~12.5 ml and ~14.4 ml, respectively, are shown for reference with bars at the top of the chromatogram. The asterisk indicates the peak containing Spike trimer. Fractions subjected to WB staining with ECL (from 2 to 14) are indicated at the bottom of the chromatogram. 4b) WB of SEC eluted fractions. Protein was revealed using an anti 6xHis tag antibody and ECL. Lane M, molecular weight markers; lane C, positive control; lane 1, input; lanes 2 to 14 correspond to respective fractions obtained from the SEC column.
Figure 5 : Analysis of ELISA performance resulting from the study of 82 sera from normal control individuals and 99 sera from COVID-19 patients. (A) Sensitivity curve (o) and specificity (x) as a function of the possible cut-off values. The vertical dashed line indicates the cut-off value with the optimized sensitivity and specificity parameters (cut-off = 3.0). (B) ROC curve analysis of ELISA, AUC is included.
Figure 6 : Antibodies anti-SARS-Cov 2 results obtained by ELISA in sera from normal control individuals and sera from COVID 19 patients IgG positive and negative by COVIDAR IgG ELISA test. Results are expressed as SDs. The cut-off value (SDs >3.0) is indicated by a dotted line and medians for each population are indicated as a full line (***p<0.0001).