Comprehensive mapping of West Nile virus (WNV)- and Japanese encephalitis virus serocomplex-specific linear B-cell epitopes from WNV non-structural protein 1

Abstract

West Nile virus (WNV) non-structural protein 1 (NS1) elicits protective immune responses during infection of animals. WNV NS1-specific antibody responses can provide the basis for serological diagnostic reagents, so the antigenic sites in NS1 that are targeted by host immune responses need to be identified and the conservation of these sites among the Japanese encephalitis virus (JEV) serocomplex members also needs to be defined. The present study describes the mapping of linear B-cell epitopes in WNV NS1. We screened eight NS1-specific mAbs and antisera (polyclonal antibodies; pAbs) from mice immunized with recombinant NS1 for reactivity against 35 partially overlapping peptides covering the entire WNV NS1. The screen using mAbs identified four WNV-specific (including Kunjin virus) epitopes, located at aa 21-36, 101-116, 191-206 and 261-276 in WNV NS1. However, using pAbs, only three WNV-specific epitopes were identified, located at positions 101-116, 191-206 and 231-246. Two of these epitopes (aa 21-36 and 261-276) had different reactivity with mAbs and pAbs. The knowledge and reagents generated in this study have potential applications in differential diagnostics and epitope-based marker vaccine development for WNV and viruses of the JEV serocomplex.

Comprehensive Mapping of Common Immunodominant Epitopes in the Eastern Equine Encephalitis Virus E2 Protein Recognized by Avian Antibody Responses

“…A set of 42 partially overlapping polypeptides, each 16 amino acids in length, covering the entirety of the EEEV E2 protein (E2-1 to E2-42, Table S1) were generated in our laboratory according to the methods previously described [35], [36]. The E2 sequence used for the experiments in this study was synthesized from an EEEV NA variant strain (GenBank accession number X63135.1) and cloned into pCI-neo, and maintained in our laboratory.…”

“…The E2 sequence used for the experiments in this study was synthesized from an EEEV NA variant strain (GenBank accession number X63135.1) and cloned into pCI-neo, and maintained in our laboratory. The plasmid pShuttle-E2 were constructed and maintained in our laboratory [35], [36].…”

Comprehensive Mapping of Common Immunodominant Epitopes in the West Nile Virus Nonstructural Protein 1 Recognized by Avian Antibody Responses

“…Purified WNV NS1 protein and 35 partially overlapping, MBP-fused polypeptides covering the entire WNV NS1 protein (NS1-1 to NS1-35) (Data S1) were generated in our laboratory as previously described [57].…”

“…The ELISA was performed as described previously, using synthesized polypeptides as coating antigen (100 ng/well) [57]. The irrelevant polypeptide (V5-Tag: GKPIPNPLLGLDST) and serum from unimmunized animals served as negative controls.…”

Identification of three novel linear B-cell epitopes on the VP5 protein of BTV16

“…The ELISA assay used has been described previously (Sun et al, 2011(Sun et al, , 2012. Briefly, microplates were sensitized at 4 8C overnight with purified His 6 -BTV16 VP5d41aa at 100 ng ml À1 .…”