ABSTRACT
Interleukin-8(IL-8)/CXCL8 is a CXC-family chemokine that attracts lymphocytes to sites of tissue damage and plays a role in the inflammatory response and wound healing. Chicken chemotactic and angiogenic factor was referred to as chCXCLi2 and has been studied as one of human CXCL8 homologue for more than 20 years. However, no monoclonal antibodies (mAbs) that specifically detect chCXCLi2 have been developed. Here, we developed and characterized mouse mAbs against chCXCLi2 to define its immunological properties. Two mouse mAbs against chCXCLi2 were generated and confirmed to display specific binding with not only recombinants, but endogenous chCXCLi2 by Western blot analysis, ELISA, and immunocytochemistry. Inhibition of chCXCLi2-induced chemotactic activity on peripheral blood lymphocytes, proliferation of chicken macrophage cells and expression of alpha smooth-muscle actin in chicken embryonic fibroblast cells by antibodies indicate that these antibodies are capable of blocking chCXCLi2 bioactivity. These chCXCLi2 mAbs will be useful reagents for future investigations of inflammation in poultry.
Subject(s)
Antibodies, Monoclonal/isolation & purification , Avian Proteins/metabolism , Chickens/immunology , Interleukin-8/metabolism , Leukocytes, Mononuclear/immunology , Animals , Antibody Specificity/immunology , Avian Proteins/genetics , Avian Proteins/immunology , Cell Proliferation , Humans , Hybridomas , Interleukin-8/genetics , Interleukin-8/immunology , Mice , Mice, Inbred BALB C , Recombinant Proteins/immunology , Sequence Homology, Amino AcidABSTRACT
Tumor necrosis factor-like ligand 1A (TL1A) is a type II transmembrane protein predominantly expressed by endothelial cells that promotes the expansion of activated T cells and regulatory T cells, modulates inflammation, and regulates the production of a wide variety of T cell cytokines. However, there have not been any mAbs which specifically detect chTL1A and define its biochemical and immunological properties. So in this study, two mouse monoclonal antibodies (mAbs) which specifically detect chicken TL1A (chTL1A) were developed and characterized. Both mAbs identified a 32 kDa Escherichia coli-derived, poly-histidine-tagged fusion protein by Western blot analysis. The mAbs identified TL1A-secreting cells in the chicken thymus, cecal tonsil, and bursa of Fabricius by immunocytochemistry, and were used to measure serum TL1A levels in normal and necrotic enteritis (NE)-afflicted chickens by antigen capture ELISA. These mAbs inhibited chTL1A-induced spleen lymphocyte proliferation, nitric oxide production by chicken macrophage cells (HD11), and blocked the cytotoxic effect of chTL1A against lymphoblastoid chicken B tumor cells (LSCC-RP9). These new mAbs that detect chTL1A will be important immune reagents for basic and applied research in poultry immunology.
