Generation and Characterization of Antibody Against Porcine Interleukin-18

Abstract

The gene-encoding mature porcine interleukin-18 (pIL-18) was amplified by PCR and cloned into a prokaryotic expression vector pET-30a(+). The resulting recombinant plasmid pET-30a-pIL-18 was transformed into Escherichia coli. Expression of recombinant pIL-18 protein was induced by 1 mM isopropyl β-D-thiogalactoside at 37°C. The purified recombinant protein was used to generate a hyperimmune antiserum in a rabbit. The anti-pIL-18 serum was evaluated for its specificity and titer through indirect enzyme-linked immunosorbent assay. The specific reactivity of the anti-pIL-18 antibody was further confirmed by Western blot and immunofluorescence assays. Moreover, the pIL-18 was able to stimulate the proliferation of pig peripheral blood mononuclear cells in vitro, indicating it may have certain biological activity.

A Monoclonal Antibody Against Transmissible Gastroenteritis Virus Generated via Immunization of a DNA Plasmid Bearing TGEV S1 Gene

“…ELISA is very convenient and simple, and we and others have used this method to detect various viruses or antibodies with great success. (9,(15)(16)(17)(18)(19)(20) In this study, we therefore used the MAb (1H4) as a primary antibody to detect TGEV. The result verified that the developed MAb 1H4 exclusively reacted with TGEV rather than a closely related porcine coronavirus PEDV, PRRSV, or avian coronvavirus IBV.…”

“…Based on the ELISA results, the spleen cells from the best immunized mice were used for fusion with SP2/0 myeloma cells and generation of hybridomas conventionally. (8,9) ELISA was adopted to test antibody production of the hybridomas, and positive hybridomas were identified and cloned at least three times. MAbs were isotyped using a mouse MAb isotyping kit (Sigma, Beijing, China).…”