Method for using light scattering in real time to directly monitor and control impurity removal in purification processes

Patent metadata

Field	Value
Publication number	US-11293930-B2
Application number	US-202117495963-A
Country	US
Kind code	B2
Filing date	Oct 7, 2021
Priority date	Mar 26, 2012
Publication date	Apr 5, 2022
Grant date	Apr 5, 2022

Abstract

The invention provides a method for controlling contaminants in biopharmaceutical purification processes by using light scattering and UV absorbance to establish a determinant. The invention makes use of multi-angle light scattering (MALS) and UV as a continuous monitoring system to provide information about the elution peak fractions in real-time instead of conventional pooling methods that rely on a predetermined percent UV peak max value to initiate the pooling process; regardless of product quality.

First claim

What is claimed is:

1. A method for purifying a recombinant protein comprising:
loading a mixture containing a recombinant protein and biological impurities on to a cation exchange chromatography resin;
eluting by a salt gradient the recombinant protein from the cation exchange chromatography resin;
measuring the intensity of light scattered by the eluate with a laser light scattering detector to obtain a light scattering (LS) signal;
measuring the absorbance of the eluate with an ultraviolet (UV) absorbance detector to obtain an absorbance signal;
calculating a ratio of the LS signal to the absorbance signal for each elution fraction to obtain a LS/UV fraction ratio;
monitoring in real time during the elution of the recombinant protein the LS/UV fraction ratio over time for each elution fraction, wherein an increase in the LS/UV fraction ratio over time indicates an increase in the levels of biological impurities in the elution pool;
terminating the elution when an increase in the LS/UV fraction ratio over time is detected; and
recovering the recombinant protein from the eluate.

2. The method of claim 1 , wherein recovering the recombinant protein in the eluate further comprises contacting the eluate with a second chromatography resin or processing step; and recovering the recombinant protein from the second chromatography resin or processing step.

3. The method of claim 1 , wherein the recombinant protein is selected from the group consisting of a human antibody, a humanized antibody, a chimeric antibody, a recombinant fusion protein, or a cytokine.

4. The method of claim 1 , wherein the recombinant protein is denosumab.

Assignees

• Amgen Inc

Inventors

• Hewig Iii Arthur C
• Phan Duke H
• Yigzaw Yinges
• Bailey Robert

Classifications

• C07K16/00 Primary

  Immunoglobulins [IG], e.g. monoclonal or polyclonal antibodies · CPC title

• C07K16/065

  Purification, fragmentation · CPC title

• G01N33/6854 Primary

  Immunoglobulins · CPC title