Fine mapping of canine parvovirus B cell epitopes.
In this report we describe the topological mapping of neutralizing domains of canine parvovirus (CPV). We obtained 11 CPV-specific monoclonal antibodies (MAbs), six of which are neutralizing. The reactivities were as determined by ELISA and Western blot (immunoblot) analysis. VP2, the most abundant protein of the CPV capsid, seemed to contain all the neutralization sites. Also, an almost full-length genomic clone of CPV was constructed in the bacterial plasmid pUC18 to enable expression of CPV proteins. All the neutralizing MAbs recognized recombinant VP2 when it was expressed as a free protein in Escherichia coli but not when expressed as a fusion protein with glutathione-S-transferase. When two large fragments containing about 85% and 67% of the C terminus of VP2 were expressed, no neutralization sites were detected. When fusion proteins containing the N terminus were expressed, two linear determinants were mapped, one between residues 1 to 10 of VP2, and the other between amino acids 11 and 23. The peptide 11 GQPAVRNERATGS 23, recognized by MAb 3C9, was synthesized chemically and checked for immunogenicity, not being able to induce neutralizing activity. Although the antibody response in rabbits to all the fusion proteins was uniformly high, the anti-CPV response was very variable. Protein from pCPVEx11, which contains a T cell epitope (peptide PKIFINLAKKKKAG) present in the VP1-specific region as well as the B cell epitopes, seemed to be the most effective in inducing virus neutralization.
INGENASA, Inmunologia y Genetica Aplicada, S.A., Madrid, Spain.
J A
JA
López de Turiso
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E
Cortés
A
A
Ranz
J
J
García
A
A
Sanz
C
C
Vela
J I
JI
Casal
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology* / Q000382:microbiology; D015153:Blotting, Western; D002213:Capsid / Q000276:immunology; D036022:Capsid Proteins; D002415:Cats; D002460:Cell Line; D003001:Cloning, Molecular; D004797:Enzyme-Linked Immunosorbent Assay; D016679:Genome, Viral; D016056:Immunodominant Epitopes; D008969:Molecular Sequence Data; D009500:Neutralization Tests; D010321:Parvoviridae / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D010455:Peptides / Q000138:chemical synthesis / Q000276:immunology; D010957:Plasmids; D011993:Recombinant Fusion Proteins / Q000276:immunology; D015183:Restriction Mapping; D015678:Viral Structural Proteins / Q000276:immunology
D000911:Antibodies, Monoclonal; D036022:Capsid Proteins; D016056:Immunodominant Epitopes; D010455:Peptides; D011993:Recombinant Fusion Proteins; D015678:Viral Structural Proteins