Hybritech Incorporated

Eli Lilly and Company

Rodney A Jue

Thomas C Furman

James P McDonough

Richard M Bartholomew

Daniel E Beidler

Novel methods for reducing the heterogeneity of secreted monoclonal antibodies are disclosed. The first method comprises incubating the heterogeneous antibodies at low pH for a length of time sufficient to convert the heterogeneous forms of antibodies into substantially homogeneous forms. Another method produces the same result using ascites fluid, while yet another method produces the same result using carboxypeptidase. The homogeneous antibodies can then be purified in high yield.

Method for the reduction of heterogeneity of monoclonal antibodies

The present invention provides a method for the separation of anti-metal chelate antibodies from non-specific proteins, including antibodies, by applying a preparation containing the anti-metal chelate antibodies to an oxo acid derivatized solid support and eluting first with an elution buffer containing sufficient salt concentration to elute non-specific proteins but not sufficient to elute the anti-metal chelate antibodies and then increasing the salt concentration of the elution solution so as to elute the anti-metal chelate antibodies. In one embodiment, the oxo acid derivatized solid support is a sulfopropyl resin. Appropriate salts include sodium phosphate, sodium chloride and sodium acetate. The method can be used to separate monoclonal or polyclonal anti-metal chelate antibodies from non-specific proteins as well as to separate bifunctional anti-metal chelate antibodies from monoclonal anti-metal chelate antibodies and other non-specific proteins. The method is also useful for separating anti-metal chelate antibody fragments bearing antigen reactive regions from non-specific proteins.

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San Diego, CA, United States of America

Rodney A Jue