Fiche publication
Date publication
janvier 2018
Journal
Analytical chemistry
Auteurs
Membres identifiés du Cancéropôle Est :
Dr CIANFERANI Sarah
,
Dr VAN DORSSELAER Alain
,
Dr CARAPITO Christine
Tous les auteurs :
Husson G, Delangle A, O'Hara J, Cianferani S, Gervais A, Van Dorsselaer A, Bracewell D, Carapito C
Lien Pubmed
Résumé
Host cell proteins (HCP) are a major class of impurities derived from recombinant protein production processes. While HCP are usually monitored by ELISA, mass spectrometry (MS)-based approaches are emerging as promising orthogonal methods. Here, we developed an original method relying on data-independent acquisition (DIA) coupling global HCP amount estimation (Top 3) and absolute quantification with isotope dilution (ID). The method named Top 3-ID-DIA was benchmarked against ELISA and a gold-standard selected reaction monitoring assay (ID-SRM). Various samples generated at different steps and conditions of the purification process, including different culture durations, harvest procedures, and purification protocols were used to compare the methods. Overall, HCP were quantified over 5 orders of magnitude and down to the sub-ppm level. The Top 3-ID-DIA strategy proved to be equivalent to the gold-standard ID-SRM in terms of sensitivity (1-10 ppm), accuracy, and precision. Moreover, 81% of the Top 3 estimations were accurate within a factor of 2 when compared to ID-SRM. Thus, our approach aggregates global HCP profiling for comprehensive process understanding with absolute quantification of key HCP within a single analysis and provides an improved support for bioprocess development and product purity assessment.
Mots clés
Animals, Antibodies, Monoclonal, analysis, CHO Cells, Cricetulus, Enzyme-Linked Immunosorbent Assay, methods, Immunoglobulin G, analysis, Mass Spectrometry, methods, Recombinant Proteins, analysis
Référence
Anal. Chem.. 2018 Jan 16;90(2):1241-1247
