title: Expression, antibody generation, and biological analysis of chicken interleukin-18.
abstract: The gene encoding mature chicken interleukin-18 (ChIL-18) was cloned into prokaryotic expression vector pET-30a(+), resulting in a recombinant plasmid pET-30a-ChIL-18. After pET-30a-ChIL-18 was transformed into Escherichia coli Rosseta, the expression of ChIL-18 induced by 1 mM IPTG at 37°C was analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The expressed fusion protein of 26 kDa was purified with a Ni-NTA affinity column and used to generate a hyperimmune antiserum in a rabbit. The specificity and titer of anti-ChIL-18 serum were analyzed by the enzyme-linked immunosorbent assay. Western blot and immunofluorescence assays indicated that the anti-ChIL-18 antibody specifically reacted with the ChIL-18 expressed from E. coli or ChIL-18-transfected eukaryotic cells. Moreover, the renatured ChIL-18 stimulated the production of nitric oxide (NO) from macrophages via eliciting the secreting of IFN-γ from lymphocytes.
authors.affiliation: College of Veterinary Medicine, Northeast Agricultural University, Xiangfang District, Harbin, China.
authors.forename: Heng
authors.initials: H
authors.lastname: Zhang
authors.forename: Guangxing
authors.initials: G
authors.lastname: Li
authors.forename: Xiaofeng
authors.initials: X
authors.lastname: Ren
mesh_terms: D000818:Animals; D000917:Antibody Formation / Q000276:immunology*; D015153:Blotting, Western; D002645:Chickens / Q000276:immunology*; D002846:Chromatography, Affinity; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004926:Escherichia coli; D019084:Fluorescent Antibody Technique, Indirect; D005822:Genetic Vectors / Q000235:genetics; D020382:Interleukin-18 / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011817:Rabbits
chemical_list: D017931:DNA Primers; D020382:Interleukin-18; D009569:Nitric Oxide