Isolation of a novel cytotoxic lymphokine (factor 2) from a human B-cell line (Karpas 160b) by preparative isoelectric focusing in the rotofor cell and chromatofocusing.
Preparative isoelectric focusing (IEF) using a Rotofor cell was used for the isolation of a biologically active form of a novel cytotoxic lymphokine (Factor 2, F2) produced by a human B-cell line (Karpas 160b). We found that F2 either purified by DE52 or extracted from 160b cells using CHAPS, after adding octyl glucoside to 0.05%, could be electrofocused in the Rotofor cell without the addition of carrier ampholytes. Better recovery and separation were also achieved in the presence of 0.5% CHAPS, 3M urea. We compared the fractionation of F2 by chromatofocusing and IEF in Rotofor. Our results also showed that F2 was separable from tumor necrosis factor and phorbol myristate acetate-associated cytotoxicity by one step IEF in Rotofor. F2 purified by DE52, S-300 and Mono Q was further purified by IEF in Rotofor.
Ni
J
J
Department of Haematology, University of Cambridge.
Karpas
A
A
D001402:B-Lymphocytes / Q000737:chemistry*; D002460:Cell Line; D002853:Chromatography, Liquid; D003602:Cytotoxicity, Immunologic / Q000502:physiology*; D006801:Humans; D007372:Interferons / Q000302:isolation & purification; D007525:Isoelectric Focusing; D007526:Isoelectric Point; D008222:Lymphokines / Q000302:isolation & purification*; D014409:Tumor Necrosis Factor-alpha / Q000302:isolation & purification
D008222:Lymphokines; D014409:Tumor Necrosis Factor-alpha; D007372:Interferons