Recombinant human interleukin-2 (IL-2) expressed as Escherichia coli was isolated as insoluble aggregates of protein (inclusion bodies) after cell breakage. IL-2 and contaminants were dissolved in 6 M-guanidinium chloride/10 mM-dithiothreitol, pH 8.5, and further purified in reduced and denatured form by gel-permeation chromatography in the same solvent. Renaturation was effected by dilution and autoxidation; IL-2 of native specific activity was isolated at over 95% purity by reversed-phase h.p.l.c.; an additional peak of reduced protein was also observed. Most losses of native IL-2 occurred on refolding, probably because of an aggregation process; concentrations around 1 microgram/ml were necessary to achieve 30% recovery. It was essential to maintain the denatured protein in reduced form before renaturation and autoxidation, which was most efficient at pH 8.5 with 1.5 microM-CuSO4. A procedure based on these observations has been used to prepare IL-2 on the 50 micrograms scale.