Characterisation of a monoclonal antibody to carp IL-1β and the development of a sensitive capture ELISA

• Published in: Fish & shellfish immunology 2002-08, Vol.13 (2), p.85-95
• Main Authors: Mathew, J.A., Guo, Y.X., Goh, K.P., Chan, J., Verburg-van Kemenade, B.M.L., Kwang, J.
• Format: Article
• Language: English
• Subjects: carp IL-1β, monoclonal antibody, proliferation assay, native cytokine
• ISSN: 1050-4648; 1095-9947
• DOI: 10.1006/fsim.2001.0383

A carp IL-1β gene was identified from a subtraction hybridisation technology based cDNA library from activated carp leucocytes. This gene was cloned into pQE vector carrying 6xHis tag and the protein was expressed. Recombinant IL-1β was used to produce hybridomas specific for carp IL-1β. Monoclonal antibodies were purified by affinity column and a sandwich ELISA for IL-1β was developed with a detection limit of 10ng of the recombinant protein. Using the capture ELISA, the presence of native IL-1β in culture supernatant of PHA-stimulated leucocytes from carp was identified, which was confirmed by SDS-PAGE and Western blot. Since IL-1β is known to stimulate proliferation of T&B cells and macrophages, its ability to stimulate proliferation of carp leucocytes was studied using tritiated thymidine. The recombinant protein was found to significantly stimulate proliferation of head kidney and spleen cells from carp.