Journal: iScience
Article Title: Selection for immune evasion in SARS-CoV-2 revealed by high-resolution epitope mapping and sequence analysis
doi: 10.1016/j.isci.2023.107394
Figure Lengend Snippet: High-density peptide arrays (HDPA) provide high-resolution antibody epitope maps across the SARS-CoV-2 proteome (A) Overview of analytical pipeline. The proteome of SARS-CoV-2 was translated into 15-mer overlapping peptides with a peptide-to-peptide overlap of 13 amino acids. The resulting individual peptides were printed in duplicates on the microarray. Sera from confirmed SARS-CoV-2-positive and -negative individuals were incubated on PEPperCHIP the HDPA. Serum antibody binding was visualized using respective fluorescently labeled secondary antibodies (anti-human IgG and anti-human IgA). Image acquisition and data quantification resulted in epitope-specific antibody profiles for SARS-CoV-2. (B) Average relative fluorescent units (RFU) profiles and peptide coverages are plotted across the SARS-CoV-2 proteome (ORF1A, ORF1B, Spike (S) protein, Envelope (E) protein, Membrane (M) glycoprotein, Nucleocapsid (N) phosphoprotein). Antibody responses to each linear 15-mer peptide were mapped across the SARS-CoV-2 proteome and average RFU calculated for each amino acid residue. The normalized positional ‘epitope coverage’ at each protein residue location is defined as the ratio of total peptides mapped to each position by the total expected peptides (see section). ‘Hotspots’ can be seen as spiked in the RFU or coverage distributions. (C) Comparison of mean RFU (log-scale) between SARS-CoV-2-positive and -negative sample groups for each viral protein. (unpaired t-test, ns: p > 0.05; ∗: p ≤ 0.05; ∗∗: p ≤ 0.01; ∗∗∗: p ≤ 0.001; ∗∗∗∗: p ≤ 0.0001).
Article Snippet: To analyze the antibody responses to SARS-CoV-2 at the epitope level we used a recently developed high-density peptide array (HDPA), the PEPperCHIP Microarray (PEPperPRINT GmbH, Germany), covering the proteome of the SARS-CoV-2 isolate Wuhan-Hu-1 as well as the four seasonal hCoVs OC43, HKU1, NL63 and 229E (see for accession numbers used).
Techniques: Microarray, Incubation, Binding Assay, Labeling, Membrane, Residue, Comparison
