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Title: Development and characterization of mouse monoclonal antibodies reactive with chicken CD80

item Lee, Sung
item Lillehoj, Hyun
item PARK, MYEONG SEON - US Department Of Agriculture (USDA)
item BALDWIN, CYNTHIA - University Of Massachusetts
item TOMPKINS, DANNIELLE - University Of Massachusetts
item WAGNER, BETTINA - Cornell University
item BABU, UMA - Food And Drug Administration(FDA)
item DEL CACHO, EMILIO - University Of Zaragoza
item MIN, WONGI - Gyeongsang National University

Submitted to: Comparative Immunology Microbiology and Infectious Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/20/2011
Publication Date: 6/23/2011
Citation: Lee, S.H., Lillehoj, H.S., Park, M., Baldwin, C., Tompkins, D., Wagner, B., Babu, U., Del Cacho, E., Min, W. 2011. Development and characterization of mouse monoclonal antibodies reactive with chicken CD80. Comparative Immunology Microbiology and Infectious Diseases. 34(3):273-279.

Interpretive Summary: Limited availability of immune reagents that can be used to assay for poultry immunity hinders the progress in disease research in poultry. In this paper, ARS scientists developed mouse monoclonal antibodies which detect chicken cytokine CD80. CD80 is an important cell surface antigen on antigen presenting cells which is necessary for T-cell activation. Ability to assess the expression and function of chicken CD80 will enhance basic and applied research on poultry immunology. These mouse antibodies detecting chicken CD80 will be commercially available through technology transfer to Kingfisher company.

Technical Abstract: CD80 is one of the ligands for CD28 and is an important co-stimulator molecule on antigen presenting cells necessary for T-cell activation. Although CD80 is well characterized in human, swine, ovine, feline, and canine species, there is no information on its chicken counterpart. This study was carried out in order to develop immunological reagents that can be used to characterize chicken CD80 (chCD80). A recombinant plasmid of chCD80/horse IgG4 was constructed and expressed in CHO cells to produce recombinant chCD80 protein. Chicken CD80 was purified from the chCD80/IgG4 fusion protein by enterokinase digestion, and used to immunize BALB/c mice, resulting in 158 hybridomas that produced monoclonal Abs (mAbs) against chCD80. Three hybridomas that produced mouse mAbs with high binding specificity for rchCD80-transfected CHO cells were selected by flow cytometric analysis for further characterization. All mAbs to chCD80 showed staining of bursa and spleen, and identified a 80 kDa protein on CD80/IgG4-transfected cells in Western blot. Immunohistochemistry of lymphoid tissues revealed that CD80-expressing cells were expressed exclusively in the bursal follicles at the outer portion of the cortex in the bursa, and throughout the red pulp and the outer boundary of the white pulp in the spleen. Addition of chCD80 mAb to Con A-stimulated spleen cells inhibited the percentage of MHCII-expressing cells and suppressed IL-2 induced lymphocyte proliferation (P < 0.05), making these CD80 specific mAbs valuable immunological reagents for basic and applied poultry immunology research.