PRODUCTION AND CHARACTERIZATION OF POLYCLONAL ANTI-CANINE INTERLEUKIN-8 ANTIBODIES

Polyclonal anti-canine interleukin-8 (cIL-8) antibodies were raised in rabbits immunized with cIL-8 expressed by E. coli. Polyclonal antibod ies were purified by affinity chromatography. In the enzyme linked imm unosorbent assay (ELISA), the resulting anti-cIL-8 antibodies showed r elatively high reactivities with cIL-8 in the fusion proteins of gluta thione-S-transferase/cIL-8 (GST/cIL-8) and maltose binding protein/cIL -8 (h4BP/cIL-8), but negligible ones with MBP. Furthermore, Western bl otting analysis using these polyclonal antibodies showed distinct band s for cIL-8, GST/cIL-8, and MBP/cIL-8. These antibodies also bound to recombinant human IL-8 (rhIL-8) in ELISA but not in Western immunoblot ting. The rhIL-8 (50-800 ng/ml) was chemoattractant for canine neutrop hils in a dose dependent manner, but the anti-cIL-8 antibodies did not show the inhibitory effect on the chemotactic activity of rhIL-8 for canine neutrophils, when tested by the chemotaxis assay using Boyden c hambers. In addition, GST/cIL-8 and rhIL-8 induced strong and rapid sh ape change responses of canine neutrophils. However, the anti-cIL-8 an tibodies inhibited shape change responses induced by GST/cIL-8 but not by rhIL-8.