CPC C07K 1/165 (2013.01) [B01D 15/3809 (2013.01); C07K 1/22 (2013.01); C07K 16/065 (2013.01); C07K 16/248 (2013.01); C07K 16/2866 (2013.01); C07K 2317/24 (2013.01)] | 8 Claims |
1. A method of purifying a mono-specific anti-IL-6 receptor antibody from a sample containing said antibody and impurities, wherein the anti-IL-6 receptor antibody is tocilizumab, the impurities comprise host cell proteins, aggregates of the anti-IL-6 receptor antibody and fragments of the anti-IL-6 receptor antibody, and the method comprises the following steps:
(a) contacting the sample containing the antibody and the impurities with a protein A chromatography material under conditions such that the antibody binds to the protein A chromatography material and at least a portion of the impurities does not bind to the protein A chromatography material;
(b) eluting the antibody from the Protein A chromatography material, in order to obtain an eluate;
(c) loading the eluate of step (b) onto a first mixed mode chromatography material under conditions such that the antibody does not bind to the first mixed mode chromatography material and at least a portion of the remaining impurities binds to the first mixed mode chromatography material, wherein the first mixed mode chromatography material is a N-benzyl-N-methyl ethanol amine ligand;
(d) recovering the flowthrough containing the antibody under conditions such that said recovered flowthrough contains a lower level of impurities than the eluate of step (b),
(e) loading the recovered flowthrough containing the antibody of step (d) onto a second mixed mode chromatography material under conditions such that the antibody does not bind to the second mixed mode chromatography material and at least a portion of the remaining impurities binds to the second mixed mode chromatography material; wherein the second mixed mode chromatography material is a fluorapatite-based ligand of ceramic fluoroapatite type I or ceramic fluoroapatite type II; and
(f) recovering the flowthrough containing the antibody under conditions such that said recovered flowthrough contains a lower level of impurities than the recovered flowthrough of step (d).
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