Determination of chromatography conditions

Patent metadata

Field	Value
Publication number	US-2016367911-A1
Application number	US-201615173215-A
Country	US
Kind code	A1
Filing date	Jun 3, 2016
Priority date	Jun 16, 2015
Publication date	Dec 22, 2016
Grant date	—

Abstract

The present invention relates to a method for the determination of chromatography conditions for the separation of a biomolecule from a liquid sample, which method comprises selecting a number of experiments using design of experiments (DoE); performing said experiments with in-line conditioning of orthogonal quality measures; and based on the results from the experiments, determining efficient chromatography conditions for said biomolecule.
The invention also relates to a system for performing the method as well as a computer program and an instrument comprising such a computer program.

First claim

1 . A method for determination of chromatography conditions for the separation of a biomolecule from a liquid sample, which method comprises
a) selecting a number of experiments using design of experiments (DoE); b) performing said experiments with in-line conditioning of at least two gradients of two selected quality measures; and, c) based on the results from said experiments, determining efficient chromatography conditions for said biomolecule;
wherein the experiments selected use at least two orthogonal quality measures.

2 . The method according to claim 1 , wherein step a) comprises i) preliminarily selecting a number of experiments using DoE and ii) adjusting said experiments such that experimental parameters of said experiments fall on at least two gradients of two selected quality measures.

3 . The method according to claim 1 , wherein at least one of said at least two gradients is a curved gradient.

4 . The method according to claim 1 , wherein at least two of said at least two gradients are curved gradients.

5 . The method according to claim 1 , wherein the quality measures are selected from the group consisting of type of chromatography resin; loading volume, wash volume, elution volume, binding buffer; elution buffer; surface determinants of the biomolecule and impurities present in the liquid sample.

6 . The method according to claim 1 , wherein the in-line conditioning comprises conditioning of a liquid sample to a predetermined pH and/or conductivity.

7 . The method according to claim 6 , which method comprises determining the relative component proportions of at least one each of: a buffer; an acid or a base; a solvent; and, optionally, a salt, for providing a liquid mixture of pre-defined pH and ionic strength, wherein the relative component proportions are determined using the equation of Debye-Hückel, wherein the ion size parameter a in the Debye-Hückel equation is determined as the weighted mean ion size of all species contributing to the ionic strength of the liquid mixture, and wherein the ionic strength of each species is used as weighting parameter.

8 . The method according to claim 7 , wherein the relative component proportions are determined using an iterative procedure.

9 . The method according to claim 8 , wherein the iterative procedure comprises:
(a) determining the relative component proportions wherein the pre-defined ionic strength of the liquid mixture is addressed to the species according to a pre-defined distribution among the species; (b) on the basis of the relative component proportions determined in the preceding step, calculating the ionic strength of each species in the mixture; (c) determining a new set of relative component proportions; taking account of the ionic strength calculated in (b), and (d) repeating the steps (b) and (c) until a predetermined convergence criteria is met.

10 . The method according to claim 9 , wherein in step (a), the pre-defined ionic strength of the liquid mixture is addressed to the salt species.

11 . The method according to claim 7 , wherein the ion size parameter a of the Debye-Hückel equation is determined as

a
=

∑

I
i



a
i

I

(
I
)

wherein in equation (I), I i is the ionic strength and a i of species i, and I the total ionic strength.

12 . The method according to claim 7 , wherein the ion size parameter a in the Debye-Hückel equation is approximated as
a= 0.5*(mass) 1/3 +shell  (II)

wherein in equation (II), the parameter “shell” is fixed at one value for a positively charged species; and at a different value for a negatively charged species.

13 . The method of claim 12 , wherein in equation (II), the parameter “shell” is fixed at a value in the range of 3.8-4.2, such as 4.0, for positively charged ionic species; and at a value in the range of 0-0.2, such as 0, for negatively charged ionic species

14 . The method according to claim 1 , wherein experiments are selected using DoE which includes multivariate analysis.

15 . The method according to claim 1 , wherein the biomolecule is an antibody, such as a monoclonal antibody, or a fragment or fusion thereof.

16 . An apparatus for the determination of chromatography conditions for the separation of a biomolecule from a liquid sample, which apparatus comprises
a) means for selecting a number of experiments including at least two gradients of two selected quality measures; and b) means for outputting to a controller for carrying out the experiments;
wherein at least two orthogonal quality measures are used and the selected experiments include in-line conditioning of buffer gradient(s).

17 . The apparatus according to claim 16 , wherein the means for selecting experiments is software capable of predicting a reduced but representative number of experiments, preferably by using DoE.

18 . The apparatus according to claim 16 , which is an instrument comprising software and optionally a robot for performing all or part of the method.

Assignees

• Ge Healthcare Bio Sciences Ab

Inventors

• Carredano Enrique Napoleon
• Rodrigo Gustav Jose

Classifications

• B01D15/424

  Elution mode · CPC title

• B01D15/362 Primary

  Cation-exchange · CPC title

• B01D15/34

  Size-selective separation, e.g. size-exclusion chromatography; Gel filtration; Permeation · CPC title

• B01D15/166

  Fluid composition conditioning, e.g. gradient · CPC title

• B01D15/3847

  Multimodal interactions · CPC title