Production and purification of bovine monocyte-derived interleukin 1.
Few studies have addressed the biological and molecular nature of bovine interleukin 1 (IL-1). In an effort to increase our understanding of the role of bovine IL-1 in bovine immunology, we investigated various parameters of its production by LPS-stimulated monocytes in vitro. Bovine monocytes isolated by our methods constitutively released IL-1 activity, as measured by the murine thymocyte IL-1 assay. Monocyte release of IL-1 activity was further augmented when the cells were incubated with 0.005-10 micrograms per ml of Escherichia coli lipopolysaccharide (LPS). The presence of 1, 5, or 10 percent heat-inactivated fetal bovine serum (FBS) enhanced LPS-stimulated bovine monocyte release of IL-1 activity as compared with monocytes cultured under serum-free conditions. We used a combination of size-exclusion and reverse-phase high-performance liquid chromatography (HPLC) to purify bovine IL-1 from serum-free monocyte culture supernatants. Size-exclusion HPLC resulted in a single peak of biological activity with an approximate molecular weight of 18,000 daltons. Further purification by reverse-phase HPLC demonstrated at least three major molecular species with IL-1 activity. Besides providing information about production of IL-1 by bovine monocytes in vitro, this study also describes a protocol to purify bovine IL-1 for future studies addressing its biological functions.
Lederer
J A
JA
Department of Pathobiological Sciences, University of Wisconsin, Madison 53706.
Czuprynski
C J
CJ
D000818:Animals; D002417:Cattle; D002851:Chromatography, High Pressure Liquid; D003470:Culture Media; D005260:Female; D066298:In Vitro Techniques; D007375:Interleukin-1 / Q000096:biosynthesis* / Q000302:isolation & purification / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology; D008970:Molecular Weight; D009000:Monocytes / Q000276:immunology* / Q000378:metabolism
D003470:Culture Media; D007375:Interleukin-1; D008070:Lipopolysaccharides