Purified activin B composition

Claim 1

A human Activin B composition, which composition has the purity (at least about 90% by weight) of Activin B produced by the method comprising: (a) diafiltering a fluid containing unpurified Activin B in 25 mM 2-(N-morpholino)ethane sulfonic acid (MES), pH 6.5, 6M urea; (b) applying the diafiltrate to a sulfopropyl-Sepharose Fast Flow column equilibrated with a buffer of 25 mM MES, pH 6.5, 6M urea; (c) adjusting the Activin-B-containing fraction from the sulfopropyl-Sepharose column with sodium chloride to 0.8M NaCl; (d) applying the NaCl-adjusted fraction to a phenyl sepharose column equilibrated with an equilibration buffer of 25 mM 3-[N-morpholino]propane sulfonic acid (MOPS), pH 7.0, 6M urea, 0.8M NaCl: (e) washing the column of step (d) sequentially with at least two column volumes of the equilibration buffer and 25 mM MOPS, pH 7.0, 6M urea; (f) eluting Activin B from the phenyl sepharose column with up to 2.5 column volumes of a mixture of about 40% ethanol and 60% 25 mM MOPS, pH 7, 6M urea; (g) applying the Activin-B-containing fraction from the phenyl sepharose column to a Sephacryl-S-300 HR column equilibrated in either (a) 25 mM MOPS, pH 7, 6M urea, or (b) 1M acetic acid, 7.1M urea, and eluting the Activin-B containing fractions from the HR column and pooling them; (h) applying the pooled fractions to a RP-HPLC column; and (i) eluting the Activin-B-containing fractions from the RP-HPLC column using a gradient mixture between water and acetonitrile, both containing 0.1% trifluoroacetic acid.