Epitope mapping of a monoclonal antibody against the Gp85 of avian leukosis virus subgroup J.
Avian leukosis virus subgroup J poses a great threat to the poultry industry in China. To reduce the economic losses, a quick method for detection of ALV-J antigen is required for diagnosis and identification of the congenitally transmitting hens. In this study, we report the production and evaluation of one monoclonal antibody (MAb) suitable for achieving these goals. The gp85 gene of avian leukosis virus subgroup J CAUHM01 China isolates was subcloned into the expression vectors pGEX-6p-1 and pET28a and successfully expressed in E. coli. After immunizing BALB/c mice with recombinant His-Jgp85 protein, splenic cells from immunized mice were fused with SP2/0 myeloma cells to produce hybridomas. We isolated and characterized one ALV-J gp85-specific MAb by determining its titer, affinity and IgG subclass. In addition, we performed epitope mapping and determined the epitope for the MAb 1E3 to be 81-92 aa of ALV-J gp85 protein (LPWDPQELDILG). Bioinformatics analysis and IFA studies revealed that this epitope is conserved among all ALV-J isolates and that this antibody could serve as a useful reagent for ALV-J detection and diagnosis.
College of Veterinary Medicine, China Agricultural University, Beijing 100193, China.
Miao
M
Sun
Duo
D
Yu
Hongfei
H
Mo
Hong
H
Cao
Chen
C
Chen
Fuyong
F
Chen
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics*; D000956:Antigens, Viral / Q000235:genetics*; D001353:Avian Leukosis / Q000175:diagnosis* / Q000821:virology; D001354:Avian Leukosis Virus / Q000235:genetics*; D015153:Blotting, Western / Q000662:veterinary; D002681:China; D019295:Computational Biology; D017931:DNA Primers / Q000235:genetics; D018604:Epitope Mapping / Q000662:veterinary*; D004926:Escherichia coli; D019084:Fluorescent Antibody Technique, Indirect / Q000662:veterinary; D005822:Genetic Vectors / Q000235:genetics; D006825:Hybridomas; D051379:Mice; D008807:Mice, Inbred BALB C; D011200:Poultry; D014759:Viral Envelope Proteins / Q000235:genetics*
D000911:Antibodies, Monoclonal; D000956:Antigens, Viral; D017931:DNA Primers; D014759:Viral Envelope Proteins; C107403:glycoprotein 85, avian leukosis virus