Genentech, Inc.

Philip M Lester

Stuart E Builder

David Reifsnyder

Roger Hart

John R Ogez

Josefine Persson

Robert L Fahrner

Jeffrey R Gorrell

Timothy N Breece

Kathlyn Pham Lazzareschi

David Peng

Deborah Ann Ansaldi

A method for purifying a desired heterologous polypeptide from microbial fermentation broth or homogenate in which it is produced and solubilized is described. This method involves adding to the broth or homogenate an effective amount of a solution of 6,9-diamino-2-ethoxyacridine lactate (ethacridine lactate) to precipitate host cell impurities under conditions wherein the majority of polypeptide remains soluble, and separating the desired polypeptide from the broth or homogenate. The broth or homogenate containing the ethacridine lactate and polypeptide is also disclosed.

Purification of microbial cell fermentation homogenates

Purification of polypeptides

A method for purifying proteins by Protein A chromatography is described which comprises removing contaminants by washing the solid phase with various intermediate wash buffers.

Protein purification

A method is disclosed for separating a polypeptide monomer from a mixture comprising dimers and/or multimers. The method comprises applying the mixture to either a cation-exchange chromatography resin or an anion-exchange chromatography resin and eluting the mixture at a gradient of about 0-1 M of an elution salt, wherein the monomer is separated from the dimers and/or multimers present in the mixture.

Separation of polypeptide monomers

Refolding of polypeptides like recombinant insulin-like growth factor

A composition is provided comprising about 0.1 to 15 mg/mL of a polypeptide in a buffer having a pH of about 7-12 comprising about 5-40% (v/v) of an alcoholic or polar aprotic solvent, about 0.2 to 3M of an alkaline earth, alkali metal, or ammonium salt, about 0.1 to 9M of a chaotropic agent, and about 0.01 to 15 .mu.M of a copper or manganese salt. The buffer is suitably used in a method for refolding improperly folded polypeptides.

Refolding of polypeptides

A method is described for isolating an exogenous polypeptide in a non-native conformation from cells, such as an aqueous fermentation broth, in which it is prepared comprising contacting the polypeptide with a chaotropic agent and preferably a reducing agent and with phase-forming species to form multiple aqueous phases, with one of the phases being enriched in the polypeptide and depleted in the biomass solids and nucleic acids originating from the cells. Preferably, the method results in two aqueous phases, with the upper phase being enriched in the polypeptide.

Aqueous multiple-phase isolation of polypeptide

Refolding of misfolded insulin-like growth factor-I

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San Lorenzo, CA, United States of America

Philip M Lester