Chromatofocusing as a tool for the characterization and partial purification of human interleukin-2.
Human interleukin-2 (IL2) has been characterized and partially purified with a sequence of chromatofocusing, gel filtration and SDS-PAGE analysis. IL2 when tested in a [3H]thymidine incorporation assay by human IL2-dependent T cells, appeared to have a MW of 25,000 as determined by Ultrogel ACA 54 gel filtration. Chromatofocusing of an 80% ammonium sulfate precipitate from crude conditioned medium yielded 4 peaks of activity corresponding to fractions of pH 7.65, 7.28, 6.72 and 6.58. Neuraminidase treatment of IL2 prior to chromatofocusing reduced its charge heterogeneity to a single peak of activity at pH 7.63. IL2 which had been treated with neuraminidase, purified by chromatofocusing, radioiodinated and further separated by gel filtration was subjected to SDS gel electrophoresis. We observed a band, migrating in the 15,000 region which only occurred in the active fractions and which we tentatively identified as IL2. These findings indicate that the purification procedure described is appropriate to the characterization and preparation of quantities of human IL2.
D002850:Chromatography, Gel; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D007376:Interleukin-2 / Q000032:analysis* / Q000302:isolation & purification; D007525:Isoelectric Focusing / Q000379:methods*; D008970:Molecular Weight; D009439:Neuraminidase / Q000494:pharmacology
D007376:Interleukin-2; D009439:Neuraminidase