Purification of proteins using hydrophobic interaction chromatography

摘要

The present invention is directed to methods for purifying a protein of interest, e.g., an antibody, from a sample comprising the protein of interest and at least one impurity, e.g., an aggregate, by employing a hydrophobic interaction chromatography (HIC) method that allows for binding of both the protein of interest and the at least one impurity under strong binding conditions. The present invention is based, at least in part, on the finding that both flow through and bind-elute techniques can be combined to achieve greater purification and recovery of a protein of interest, e.g., an antibody, under isocratic wash conditions and strong binding conditions.

主权项

1. A method for producing a preparation comprising a protein of interest and having a reduced level of at least one impurity, said method comprising:
(a) contacting a sample comprising the protein of interest and at least one impurity, to a hydrophobic interaction chromatography (HIC) media, in the presence of a load buffer such that (i) a portion of the protein of interest binds to the HIC media at a Kp of greater than 20 and (ii) a substantial portion of the at least one impurity binds to the HIC media; (b) collecting a flow through fraction comprising the protein of interest unbound to the HIC media; (c) washing the HIC media with a wash buffer such that a substantial portion of the protein of interest bound to the HIC media is released from the media, wherein the salt concentration and/or the pH of the wash buffer are within 20% of the salt concentration and/or pH of the load buffer; and (d) collecting a wash fraction comprising the protein of interest released from the HIC media, wherein each of the flow through and wash fractions comprise the protein of interest and have a reduced level of the at least one impurity.