Purification and characterization of a cytotoxic factor produced by a mouse macrophage hybridoma.
A mouse macrophage cytotoxic factor was purified to homogeneity from the serum-free culture supernatant of a mouse macrophage hybridoma clone, N/P-7-1, stimulated with lipopolysaccharide by gel filtration, affinity chromatography, anion-exchange chromatography, and polyacrylamide gel electrophoresis. The purified material was judged to be homogeneous as to the criteria of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and has a relative molecular mass of 17,500, as determined by SDS-PAGE, or 55,000, as determined by gel filtration on columns of both Sephacryl S-200 and TSK G3000SW. It has an isoelectric point of 5.0, and is trypsin sensitive, stable at 56 degrees C and labile at pH less than 6. The cytotoxic activity of the purified factor could not be inhibited by various sugars and lectins. The production of the factor from N/P-7-1 triggered by macrophage-activating factor for cytotoxicity, but not by mouse recombinant gamma-interferon. The factor should be synthesized after lipopolysaccharide stimulation because treatment of N/P-7-1 cells with a metabolic inhibitor, emetine or actinomycin D, prevents the production.
D000818:Animals; D002241:Carbohydrates / Q000494:pharmacology; D002845:Chromatography; D003603:Cytotoxins / Q000302:isolation & purification*; D004586:Electrophoresis; D006358:Hot Temperature; D006825:Hybridomas / Q000276:immunology*; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D037102:Lectins / Q000494:pharmacology; D008262:Macrophage Activation; D008264:Macrophages / Q000276:immunology*; D051379:Mice; D008970:Molecular Weight; D011480:Protease Inhibitors / Q000494:pharmacology
D002241:Carbohydrates; D003603:Cytotoxins; D037102:Lectins; D011480:Protease Inhibitors