Identification of a Novel Conserved B Cell Epitope in the N Protein of Equine Arteritis Virus (Bucyrus Strain).
Abstract: The nucleocapsid (N) protein is the most conserved structural protein in equine arteritis virus (EAV). This study aimed to identify the minimal conserved B cell epitope on the EAV N protein. The purified N protein was used to immunize mice for preparing monoclonal antibody (mAb). The reactivity of mAb was evaluated by Western blot and immunofluorescence assay. Moreover, 11 overlapping peptides (named MBP-N1 to MBP-N11) were designed to localize the linear antigenic epitope within the N protein. The peptides were identified by indirect enzyme-linked immunosorbent assay (ELISA) and Western blot. The minimal conserved B cell epitope on the EAV N protein was identified. The homology analysis was also performed. An EAV N-reactive mAb was selected and designated as 1C11. Indirect ELISA results showed that overlapping domain between MBP-N10 and MBP-N11 was recognized by the mAb 1C11. Furthermore, the indirect ELISA and Western blot showed that (101)QRKVAP(106) was the minimal linear epitope of the EAV N protein. The homology analysis showed that the identified epitope was conserved among all EAV strains analyzed in this work, with the exception of the ARVAC. One EAV N-specific mAb (1C11) was developed, and a minimal linear peptide epitope ((101)QRKVAP(106)) within the N protein was identified.
Publication Date: 2015-09-04 PubMed ID: 26331346DOI: 10.1089/vim.2015.0018Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
- Amino Acid Sequence
- Antibodies
- B cells
- Diagnosis
- Disease
- Disease Diagnosis
- Enzyme-Linked Immunosorbent Assay (ELISA)
- Equine Health
- Equine Viral Arteritis
- Immunization
- Immunofluorescence Assay
- Immunoglobulins
- Infection
- Infectious Disease
- Monoclonal Antibodies
- Peptides
- Veterinary Medicine
- Veterinary Research
- Virology
- Virus
- Western Blot
Summary
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This research examines the most conserved protein, the nucleocapsid or N protein, of the equine arteritis virus (EAV) and identifies the smallest shared B cell epitope on it. This was achieved by immunizing mice with the N protein, generating monoclonal antibodies and using a variety of tests to identify the minimal conserved epitope.
Methods
- The purified N protein from EAV was used to immunize mice to generate monoclonal antibodies (mAbs).
- The reactivity of the produced mAbs was evaluated using Western blot and immunofluorescence assay. This step aimed to confirm that the antibodies were able to detect and interact with the N protein.
- To further understand and localize the epitope (area of the antigen recognized by the antibody) within the N protein, the researchers designed 11 overlapping peptides, named MBP-N1 to MBPB-N11.
- Indirect enzyme-linked immunosorbent assay (ELISA) and Western blot were used to identify the peptides recognized by the generated mAbs.
Results
- The minimal conserved B cell epitope on the EAV N protein, or the smallest shared protein sequence that is recognized by the B cells of the immune system, was identified successfully.
- An EAV N-reactive mAb, designated as 1C11, was selected out of those produced. Indirect ELISA results showed that the mAb 1C11 recognized the overlapping domain between MBP-N10 and MBP-N11.
- Further studies using indirect ELISA and Western blot revealed that the minimal linear epitope of the EAV N protein was (101)QRKVAP(106), a sequence of six amino acids.
- The researchers compared this epitope with other EAV strains and found it was conserved, present in all of them except one strain known as ARVAC.
Conclusion
- The researchers were successful in creating an EAV N-specific mAb (1C11) and identified a minimal linear peptide epitope ((101)QRKVAP(106)) present within the N protein.
- This discovery is significant, as it can help in designing diagnostic tests for EAV based on this specific antibody interaction, or potentially in treatment strategies targeting this conserved epitope.
Cite This Article
APA
Chen J, Guo X, Li L.
(2015).
Identification of a Novel Conserved B Cell Epitope in the N Protein of Equine Arteritis Virus (Bucyrus Strain).
Viral Immunol, 28(7), 391-396.
https://doi.org/10.1089/vim.2015.0018 Publication
Researcher Affiliations
- 1 Department of Gynecology, The Third Affiliated Hospital of Harbin Medical University , Harbin, China .
- 2 Department of General Surgery, The Fourth Affiliated Hospital of Harbin Medical University , Harbin, China .
- 1 Department of Gynecology, The Third Affiliated Hospital of Harbin Medical University , Harbin, China .
MeSH Terms
- Animals
- Antibodies, Monoclonal / immunology
- Antibodies, Monoclonal / isolation & purification
- Antibodies, Viral / immunology
- Antibodies, Viral / isolation & purification
- Blotting, Western
- Conserved Sequence
- Enzyme-Linked Immunosorbent Assay
- Epitope Mapping
- Epitopes, B-Lymphocyte / genetics
- Epitopes, B-Lymphocyte / immunology
- Equartevirus / genetics
- Equartevirus / immunology
- Female
- Fluorescent Antibody Technique
- Mice, Inbred BALB C
- Nucleocapsid Proteins / genetics
- Nucleocapsid Proteins / immunology
- Sequence Homology, Amino Acid
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