Development of a sandwich ELISA for the detection of chicken colony-stimulating factor 1

Macrophage colony-stimulating factor-1 (M-CSF-1 or CSF-1) is a hematopoietic growth factor that stimulates the survival, proliferation, and differentiation of the mononuclear phagocyte lineage and is involved in bone metabolism, fertility, pregnancy, inflammatory processes, and homeostasis. CSF-1-activated macrophages display unique features, such as distinguishable cell surface antigens, enhanced Fc-γ-receptor-mediated phagocytosis, intensified reactive oxygen species activity, enhanced proliferation, and enhanced chemotaxis. Five mouse monoclonal antibodies (mAbs) for the detection of chicken CSF-1 were developed and characterized using western blot, indirect ELISA, and in vitro functional assays. One of the anti-chCSF-1 mAbs, 8A12, showed neutralization of chicken macrophage cell line (HD11) proliferation and CSF-induced nitric oxide release, whereas mAb 1G4 inhibited the phagocytosis of fluorescent-labeled E. coli by HD11 cells in vitro. For the quantitative assessment of native chCSF-1 in biological samples from chickens, a sensitive sandwich ELISA was developed using the best capture and detection pair of mAbs that were selected from newly developed anti-chCSF-1 mAbs. Chickens that were challenged with Eimeria acervulina, E. maxima, and E. tenella showed a steady increase in the circulating levels of serum CSF-1, starting from day 1 to 7 postchallenge reaching their peak levels at day 10 postchallenge infection. The CSF-1 synthesis induced by 3 different species of Eimeria was quite similar, even though these they are reported to be phenotypically and immunologically different. Therefore, this mAb-based sandwich ELISA will be a valuable tool for the detection of CSF-1 production during various poultry infections, and these new anti-chCSF-1 mAbs will facilitate the fundamental and applied research related to CSF-1 function in normal and disease states in chickens.