Establishment of swine interleukin-6 sandwich ELISA.

Abstract

We established a sandwich enzyme-linked immunosorbent assay (ELISA) for swine interleukin-6 (SwIL-6), which was applied for detection of SwIL-6 in vitro and in vivo. Anti-SwIL-6 rabbit- and goat-polyclonal antibodies, and monoclonal antibody (mAb) were prepared, conforming that all of the antibodies were reactive with recombinant SwIL-6 by Western blotting and indirect ELISA. A sandwich ELISA was developed using the mAb as a capture antibody and biotinylated goat-polyclonal antibody as a detection antibody. The detection limit of the sandwich ELISA for rSwIL-6 was 49pg/ml and did not show cross-reactivity with swine IL-1b, IL-4, IL-8, IL-18, IL-12, and IFN-g. Using the ELISA, SwIL-6 was detected in culture medium of the monocytes stimulated with PHA-P and PMA, and the plasma or the bronchoalveolar lavage fluid (BALF) of pigs experimentally infected with Actinobacillus pleuropneumoniae or Mycoplasma hyopneumoniae. This ELISA for SwIL-6 may be useful for understanding the role of this cytokine in various swine diseases.