Method for the production of domain antibodies

摘要

The present invention relates to a method for producing a domain antibody in a yeast, wherein the formation of disulfide bridges in the domain antibody is promoted. The method encompasses the addition of oxidizing agents, preferably oxidizing metal ions, preferably one or more selected from Cu2+, Fe2+, Fe3+ and Zn2+.

主权项

1. Method for producing a domain antibody in a yeast, comprising
a) culturing said yeast in culture medium under conditions that are such that said yeast will multiply; b) maintaining said yeast under conditions that are such that said yeast expresses and/or produces the domain antibody; followed by c) isolating and/or purifying the secreted domain antibody from the medium;wherein conditions are applied that promote the formation of disulfide bridges in domain antibodies selected from one or more of the following
i) addition of Cu2+ to a final concentration of higher than 100 μM during the culturing phase; ii) addition of Cu2+ to a final concentration higher than 100 μM during the production phase of the domain antibody; iii) treating the culture broth after fermentation with Cu2+ to a final concentration higher than 100 μM and with a treatment time from 0.5 h to overnight treatment; iv) treating the culture broth after fermentation with Cu2+ to a final concentration higher than 100 μM at pH of approximately 6, 7, 8 or 9 (each ±0.5); and v) addition of Cu2+ to a final concentration higher than 100 μM to the supernatant comprising the domain antibody after removal of the host, or at any step after purification of the domain antibody, or to the purified domain antibody.