Direct analysis of mAb aggregates in mammalian cell culture supernatant

BMC Biotechnology

Friedemann Hesse¹  Karen Schwab¹  Albert J Paul¹ 

[1]Institute of Applied Biotechnology (IAB), Biberach University of Applied Sciences, Biberach, 88400, Germany

关键词: CHO cells;    Mammalian cell culture;    Monoclonal antibodies;    Protein aggregation;   

Others  :  1084353
DOI  :  10.1186/s12896-014-0099-3

 received in 2014-06-06, accepted in 2014-11-14,  发布年份 2014

【 摘 要 】

Background

Protein aggregation during monoclonal antibody (mAb) production can occur in upstream and downstream processing (DSP). Current methods to determine aggregate formation during cell culture include size exclusion chromatography (SEC) with a previous affinity chromatography step in order to remove disturbing cell culture components. The pre-purification step itself can already influence protein aggregation and therefore does not necessarily reflect the real aggregate content present in cell culture. To analyze mAb aggregate formation directly in the supernatant of Chinese hamster ovary (CHO) cell culture, we established a protocol, which allows aggregate quantification using SEC, without a falsifying pre-purification step.

Results

The use of a 3 μm silica SEC column or a SEC column tailored for mAb aggregate analysis allows the separation of mAb monomer and aggregates from disturbing cell culture components, which enables aggregate determination directly in the supernatant. Antibody aggregate analysis of a mAb-producing CHO DG44 cell line demonstrated the feasibility of the method. Astonishingly, the supernatant of the CHO cells consisted of over 75% mAb dimer and larger oligomers, representing a substantially higher aggregate content than reported in literature so far.

Conclusion

This study highlights that aggregate quantification directly in the cell culture supernatant using appropriate SEC columns with suitable mAb aggregate standards is feasible without falsification by previous affinity chromatography. Moreover, our results indicate that aggregate formation should be addressed directly in the cell culture and is not only a problem in DSP.

【 授权许可】

   
2014 Paul et al.; licensee BioMed Central Ltd.

【 图 表 】

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