Stirred tank reactor and method

• US 9,803,165 B2
• Filed: 09/12/2012
• Issued: 10/31/2017
• Est. Priority Date: 12/16/2008
• Status: Active Grant

• Current Assignee: EMD Millipore Corporation (Merck & Co., Inc.)
• Original Assignee: EMD Millipore Corporation (Merck & Co., Inc.)
• Inventors: Moya, Wilson, Dupont, Alison
• Primary Examiner(s): Brown, Melanie Y
• Assistant Examiner(s): Moerschell, Richard
• Application Number:
  US13/610,954
• Publication Number:
  US 20130005950A1
• Time in Patent Office: 1,875 Days
• Field of Search: None
• CPC Class Codes: C12M 23/20 Material Coatings immunocoatings C12M25/00
• CPC Class Codes: C12M 23/28 disposable or single use
• CPC Class Codes: C12M 27/00 Means for mixing, agitating or circulating fluids in the vessel by introduction of gas C12M29/06, C12M29/14, mixing in general or mixers per se B01F; mixing in apparatus for chemical or physical processes B01J
• CPC Class Codes: C12M 47/12 Purification C12M47/04 takes precedence

Abstract

Container for sample preparation or processing, such as biomass culturing or processing, and optionally sample purification. In certain embodiments, the reactor is a bioreactor that includes a stirred cell device that simulates a tangential flow filter to reduce or eliminate clogging that can be caused by the solids generated. In certain embodiments, the solids comprise a precipitate or floc or beads, such as one that includes a polymer that binds the biomolecule(s) of interest, and impurities. In its method aspects, embodiments disclosed herein include purification and isolation of biomolecules of interest derived from cell culture fluids. The methods include carrying out sample preparation or processing in a container, culturing a biomass; generating solids by precipitating or flocculating a biomolecule of interest from the cultured broth; preventing the solids from settling in the container by agitation; and purification, such as by eluting the biomolecule of interest and filtering the same.

Claims

1. A method for purifying a biomolecule selected from the group consisting of proteins and antibodies, from a mixture containing impurities selected from the group consisting of cells, cell debris, host cell proteins and DNA, comprising:

a. providing an assembly comprising providing a first container having an interior space, a first base sealingly affixed to said first container and supporting at least one membrane sealed to said first base for filtering said mixture;

an outlet in said base, a second container in fluid communication with the outlet of said first base, and a second base sealingly affixed to said second container, said second base comprising a second membrane and a supporting surface comprising grooves for fluid flow;

b. providing the mixture at a set of conditions,

c. adding one or more polymers, soluble in said mixture under the set of conditions and capable of reversibly and selectively binding to the biomolecule, said one or more polymers selected from the group consisting of poly(N-vinyl caprolactam), poly(N-acryloylpiperidine), poly(N-vinylisobutyramide), poly(N-isopropylacrylamide), poly(N,N′

-diethylacrylamide), poly(N-acryloyl-N-alkylpiperazine), hydroxyalkylcellulose, copolymers of acrylic acid and methacrylic acid, and polymers and copolymers of 2 or 4-vinylpyridine and chitosan with either a ligand or functional group attached to it,

d. mixing the one or more solubilized polymers throughout the mixture;

e. precipitating the one or more polymers and bound biomolecule out of solution by changing the set of conditions in the mixture;

f. washing said precipitate by contacting said precipitate with a wash solution and filtering the supernatant through said first membrane, and

g. recovering the bound biomolecule from the polymer and filtering the biomolecule through said second membrane.