Top-down LC-MS quantitation of intact denatured and native monoclonal antibodies in biological samples

The requirements for developing antibody biotherapeutics benefit from understanding the nature and relevant aspects of the entire molecule. The method presented herein employs on-line multidimensional LC-quadrupole time-of-flight (QTOF)-MS for the quantitative determination of an antibody isolated f...

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Bibliographic Details
Published in:Bioanalysis 2018-07, Vol.10 (13), p.1039-1054
Main Authors: Zhang, Liyun, Vasicek, Lisa A, Hsieh, SuChun, Zhang, Shuli, Bateman, Kevin P, Henion, Jack
Format: Article
Language:English
Subjects:
antibody
biotherapeutic quantitation
denatured and native protein conformation
immunocapture
intact top-down protein quantitation
multidimensional LC-MS
non-specific binding
NSB
top-down QTOF LC/MS
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Summary:The requirements for developing antibody biotherapeutics benefit from understanding the nature and relevant aspects of the entire molecule. The method presented herein employs on-line multidimensional LC-quadrupole time-of-flight (QTOF)-MS for the quantitative determination of an antibody isolated from biological samples while maintaining the intact native biologically active conformation of the antibody. Following method optimization for a model antibody, an incurred biotherapeutic in cynomologus monkey was quantified in its intact top-down native conformation. A partial method validation demonstrated acceptable precision and accuracy although improved sensitivity requires further studies. An on-line multidimensional LC-MS approach presents a proof-of-principle example for quantifying an intact, native antibody isolated from an incurred biological sample via immunoaffinity techniques coupled with top-down QTOF LC-MS bioanalysis.
ISSN:1757-6180
1757-6199
DOI:10.4155/bio-2017-0282