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Expression, purification and monoclonal antibodies preparation of recombinant equine mature interleukin-18

Author:
Tong, Tiegang, Bai, Yu, Liu, Guangliang, Wang, Qun, Zhang, Weijun, Xiao, Yihong, Xu, Shulan, Liu, Nihong, Yang, Tao, Wu, Donglai
Source:
Veterinary immunology and immunopathology 2010 v.136 no.3-4 pp. 194-200
ISSN:
0165-2427
Subject:
horses, interleukins, recombinant proteins, synergism, interferons, interleukin-12, gene expression, in vitro culture, purification, prokaryotic cells, Escherichia coli, affinity chromatography, protective effect, immunization, immunomodulators, monoclonal antibodies, vaccination, vaccine development
Abstract:
IL-18 is a cytokine originally discovered as an important modulator of immune responses and subsequently shown to be pleiotropic. In this report, we expressed the recombinant equine mature interleukin-18 (rEMIL-18) in E. coli and purified it by nickel affinity gel column chromatography. Purified rEMIL-18 had biological activity commensurate with recombinant human IL-18, as determined by its synergistic effect with recombinant human IL-12 (rhIL-12) on the induction of IFN-γ gene expression in equine peripheral blood mononuclear cells (PBMC). Following intraperitoneal (i.p.) immunization of BALB/c mice with rEMIL-18, nine monoclonal antibodies (mAbs) against equine interleukin-18 (EIL-18) were obtained and characterized. These mAbs recognized different epitopes on equine mature interleukin-18 (EMIL-18) protein based on their reactivity with two peptides containing different amino acid sequences and one of these mAbs has neutralization activity against EIL-18 in an IFN-γ-induction assay.
Agid:
788723