Method for identification and purification of multi-specific polypeptides

摘要

The document pertains to a method for the purification of a ternary mixture of dimeric antibodies of the type AA, AB, BB, characterized in that for the separation of the three components and in particular for the isolation of the multi-specific fraction AB multicolumn counter current solvent gradient purification chromatography with a stationary phase load of more than 1 mg antibody mixture per milliliter stationary phase is used. It furthermore relates to a method for the identification of in particular bispecific antibody systems, which are particularly suitable for the application of such a purification method.

主权项

1. A method for the identification of a purifyable multi-specific polypeptide species (AiBj) which is a multimer consisting of at least two different specificity polypeptide chains (Ai, Bj), the method comprising the following steps:
(A) generation of a library of nucleic acids for each of the different specificity polypeptide chains (A, B) forming the multimer, and narrowing the library size through antigen binding and/or activity assays; (B) individually expressing a sufficient amount of polypeptide chains (A1, A2, A3, . . . An; B1, B2, B3, . . . Bm) with each individual library member, and forming homomultimers therefrom; (C) using chromatography, for the determination of an analytically discriminative parameter, for each of the homomultimers; (D) selecting at least one pair (AiAi, BjBj) of homomultimers of different specificity polypeptide chains which are analytically sufficiently discriminated; (E) expressing the polypeptide chains from the library of step (A) corresponding to the homomultimers identified in step (D) in host cells such that a ternary mixture of homomultimers (AiAi; BjBj) and heteromultimers (AiBj) is produced by culture of the host cells; and (F) purifying the resulting ternary mixture by using chromatography.