Figure 4. N protein (p40) peptide microarray–based epitope mapping of variegated squirrel bornavirus 1 from patient who died of limbic encephalitis, Germany, 2013. A) The N protein–based peptide microarray chip consists of 8 identical arrays composed of 360 15-mer peptides with an offset of 1 aa. Each subarray was bordered by biotin spots (green). B) Representative single-channel readouts from 1 subarray in 16-bit pseudocolor is given for the protein A–purified patient cerebrospinal fluid sample. Signals were classified positive if they appeared in each of the 8 subarrays. The detected signal is surrounded by an orange box and labeled by the corresponding spot number. C) The alanine scan of spot 116 showed highest antibody binding for the original sequence and less bound antibodies after substitution of the indicated amino acids by alanine. D) Mapping of antibody target sequences onto the tetrameric P40 nucleoprotein 3D-structure (PDB:1N93). Monomers are depicted in gray and gold, amino acids necessary for antibody binding (red) and inconsequential (pink) related to spot 116. Potential binding sites of antibodies for spot 116 are shown in the lower panels as red (1 monomer) and blue (spanning 2 adjacent monomers and forming a binding pocket) overlay.