Chromatography method
AssigneeGE Healthcare BioProcess R&D AB
InventorsJean-Luc Maloisel, Ola Lind, Bjorn Noren, Ronnie Palmgren
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Abstract
The present invention relates to a method for removal of large contaminants, such as virus, in a chromatographic process for purification of a target molecule, preferably monoclonal antibodies, mAbs, by using a specifically designed chromatographic bead having a thin outer layer and a core functionalized with a ligand adsorbing the mAbs or parts thereof.
Claims20
A method of making affinity chromatography beads suitable for purification of immunoglobulin containing proteins or parts thereof, from contaminants that are larger than immunoglobulin containing proteins, the method comprising:
- a) providing allylated chromatography beads having an allylated shell layer and an allylated inner core wrapped around by the allylated shell layer;
- b) inactivating the allylated shell layer of the beads to form a lid of hydrolyzed allyl groups having a thickness of 1-8 μm;
- c) activating the allylated inner core of the beads while leaving the lid intact to form activated beads;
- and
- d) coupling the activated beads with an affinity ligand to form the affinity chromatography beads;
- wherein the lid and the inner core of the beads having a defined pore size corresponding to KD of 0.1 to 1.
A method of making affinity chromatography beads suitable for purification of immunoglobulin containing proteins or parts thereof, from contaminants that are larger than immunoglobulin containing proteins, the method comprising:
- a) providing allylated chromatography beads having an allylated shell layer and an allylated inner core wrapped around by the allylated shell layer;
- b) inactivating the allylated shell layer of the beads to form a lid of hydrolyzed allyl groups having a thickness of 3-5 μm;
- c) activating the allylated inner core of the beads while leaving the lid intact to form activated beads;
- and
- d) coupling the activated beads with an affinity ligand to form the affinity chromatography beads;
- wherein the average diameter size of the chromatography bead is 250-450 μm.
- wherein the lid and the inner core of the beads having a defined pore size corresponding to KD of about 0.3 to 0.7,
- wherein the porosity of the lid and core is 0.60 to 0.85, and
- wherein the lid and core are made of agarose.
Affinity chromatography beads suitable for purification of immunoglobulin containing proteins or parts thereof, from contaminants that are larger than immunoglobulin containing proteins, manufactured by:
- a) providing allylated chromatography beads having an allylated shell layer and an allylated inner core wrapped around by the allylated shell layer;
- b) inactivating the allylated shell layer of the beads to form a lid of hydrolyzed allyl groups having a thickness of 3-5 μm;
- c) activating the allylated inner core of the beads while leaving the lid intact to form activated beads;
- and
- d) coupling the activated beads with a Z4 ligand to form the affinity chromatography beads;
- wherein the average diameter size of the chromatography bead is 250-450 μm.
- wherein the lid and the inner core of the beads having a defined pore size corresponding to KD of about 0.3 to 0.7,
- wherein the porosity of the lid and core is 0.60 to 0.85, and
- wherein the lid and core are made of agarose.
Dependencies are inferred from the claim text.