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Semiautomated pH gradient ion-exchange chromatography of monoclonal antibody charge variants

Citation

Talebi, M and Shellie, RA and Hilder, EF and Lacher, NA and Haddad, PR, Semiautomated pH gradient ion-exchange chromatography of monoclonal antibody charge variants, Analytical Chemistry, 86, (19) pp. 9794-9799. ISSN 0003-2700 (2014) [Refereed Article]

Copyright Statement

Copyright 2014 American Chemical Society

DOI: doi:10.1021/ac502372r

Abstract

A new approach using a chromatography system equipped with isocratic pumps and an electrolytic eluent generator (EG) is introduced, replacing external pH gradient delivery using conventional gradient systems, in which bottled buffers with preadjusted pH are mixed using a gradient pump. The EG is capable of generating high purity base or acid required for online preparation of the buffer at the point of use, utilizing deionized water as the only carrier stream. Typically, the buffer was generated from online titration of a reagent composed of low molecular weight amines. The reagent was delivered isocratically into a static mixing tee, where it was titrated to the required pH with electrolytically generated base or acid. The required pH gradient was thus conveniently generated by electrically controlling the concentration of titrant. Also, since the pH was adjusted at the point of use, this approach offered enhanced throughput in terms of eluent preparation time and labor, and with a more reproducible pH profile. The performance of the system was demonstrated by running pH gradients ranging from pH 8.2 to 10.9 on a polymer monolith cation-exchange column for high throughput profiling of charge heterogeneity of intact, basic therapeutic monoclonal antibodies. A high degree of flexibility in modulating the key parameters of the pH gradient, including the buffer concentration, the pH gradient slope and the operating pH range was demonstrated. This enabled fine-tuning of the separation conditions for each individual antibody in order to enhance the chromatographic resolution.

Item Details

Item Type:Refereed Article
Keywords:ion chromatography, pharmaceutical analysis
Research Division:Chemical Sciences
Research Group:Analytical Chemistry
Research Field:Separation Science
Objective Division:Manufacturing
Objective Group:Instrumentation
Objective Field:Scientific Instruments
Author:Talebi, M (Dr Mohammad Talebi)
Author:Shellie, RA (Associate Professor Robert Shellie)
Author:Hilder, EF (Professor Emily Hilder)
Author:Haddad, PR (Professor Paul Haddad)
ID Code:98041
Year Published:2014
Funding Support:Australian Research Council (DP0663781)
Web of Science® Times Cited:6
Deposited By:Chemistry
Deposited On:2015-01-29
Last Modified:2015-10-30
Downloads:0

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