Insight into purification of monoclonal antibodies in industrial columns via studies of Protein A binding capacity by in situ ATR-FTIR spectroscopy

Therapeutic monoclonal antibodies (mAbs) are effective treatments for a range of cancers and other serious diseases, however mAb treatments cost on average ∼$100 000 per year per patient, limiting their use. Currently, industry favours Protein A affinity chromatography (PrAc) as the key step in down...

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Bibliographic Details
Published in:Analyst (London) 2021-08, Vol.146 (16), p.5177-5185
Main Authors: Beattie, James W., Rowland-Jones, Ruth C., Farys, Monika, Tran, Richard, Kazarian, Sergei G., Byrne, Bernadette
Format: Article
Language:English
Subjects:
Binding
Chemistry
Denaturation
Fourier transforms
In situ leaching
Infrared analysis
Infrared reflection
Infrared spectroscopy
Leaching
Ligands
Monoclonal antibodies
Production costs
Protein A
Proteins
Resins
Side inlets
Spectroscopic analysis
Spectrum analysis
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Summary:Therapeutic monoclonal antibodies (mAbs) are effective treatments for a range of cancers and other serious diseases, however mAb treatments cost on average ∼$100 000 per year per patient, limiting their use. Currently, industry favours Protein A affinity chromatography (PrAc) as the key step in downstream processing of mAbs. This step, although highly efficient, represents a significant mAb production cost. Fouling of the Protein A column and Protein A ligand leaching contribute to the cost of mAb production by shortening the life span of the resin. In this study, we assessed the performance of used PrAc resin recovered from the middle inlet, center and outlet as well as the side inlet of a pilot-scale industrial column. We used a combination of static binding capacity (SBC) analysis and Attenuated Total Reflection-Fourier Transform Infrared (ATR-FTIR) spectroscopy to explore the used resin samples. SBC analysis demonstrated that resin from the inlet of the column had lower binding capacity than resin from the column outlet. ATR-FTIR spectroscopy with PLS (partial least square) analysis confirmed the results obtained from SBC analysis. Importantly, in situ ATR-FTIR spectroscopy also allowed both measurement of the concentration and assessment of the conformational state of the bound Protein A. Our results reveal that PrAc resin degradation after use is dependent on column location and that neither Protein A ligand leaching nor denaturation are responsible for binding capacity loss.
ISSN:0003-2654
1364-5528
DOI:10.1039/d1an00985k