Static coatings for the analysis of intact monoclonal antibody drugs by capillary zone electrophoresis

This work reports on the investigation of the optimal conditions associated with good isoform resolution, minimized adsorption, and acceptable repeatability for the analysis of intact monoclonal antibodies. To this end, various static capillary coatings were evaluated, and major trends were observed...

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Bibliographic Details
Published in:Electrophoresis 2013-09, Vol.34 (18), p.2718-2724
Main Authors: Gassner, Anne-Laure, Rudaz, Serge, Schappler, Julie
Format: Article
Language:English
Subjects:
Acetates - chemistry
Acetic Acid - chemistry
Adsorption
Antibodies, Monoclonal - analysis
Antibodies, Monoclonal - chemistry
Capillarity
Coating
Coatings
Criteria
CZE
Electroosmosis
Electrophoresis
Electrophoresis, Capillary - instrumentation
Electrophoresis, Capillary - methods
Intact protein
Monoclonal antibodies
Monoclonal antibody
Recovery
Reproducibility of Results
Static Electricity
Surface chemistry
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Summary:This work reports on the investigation of the optimal conditions associated with good isoform resolution, minimized adsorption, and acceptable repeatability for the analysis of intact monoclonal antibodies. To this end, various static capillary coatings were evaluated, and major trends were observed. For a positively charged coating, a low EOF mobility value was the main criterion for satisfactory resolution. For neutral coatings, adsorption was the main concern. If the coating does not fully cover the surface silanols, electrostatic interactions tend to decrease protein recovery through irreversible adsorption. The influence of the BGE pH was also investigated, as the resolution of monoclonal antibody isoforms is driven by their charge difference. The nature and concentration of the BGE were proven to be important, as an ampholytic compound such as ε‐aminocaproic acid was found to decrease adsorption compared with ammonium acetate. With the optimal conditions determined during method development, a complete investigation according to four criteria, i.e. migration time, separation efficiency, EOF mobility, and protein recovery, did not present evidence of any adsorption. Finally, the repeatability and intermediate precision were assessed, and low variability was demonstrated with an RSD less than 1.3% for the main peak migration time.
ISSN:0173-0835
1522-2683
DOI:10.1002/elps.201300070