title: Proteome Analysis Is a Valuable Tool to Monitor Antigen Expression during Upstream Processing of Whole-Cell Pertussis Vaccines.
abstract: Physicochemical and immunochemical assays were applied to substantiate the relation between upstream processing and the quality of whole-cell pertussis vaccines. Bordetella pertussis bacteria were cultured on a chemically defined medium using a continuous cultivation process in stirred tank reactors to obtain uniform protein expression. Continuous culture favors the consistent production of proteins known as virulence factors. Magnesium sulfate was added during the steady state of the culture in order to diminish the expression of virulence proteins. Changes in gene expression and antigen composition were measured by microarrays, mass spectrometry and ELISA. Transcriptome and proteome data revealed high similarity between the biological triplicates demonstrating consistent cultivation of B. pertussis. The addition of magnesium sulfate resulted in an instant downregulation of the virulence genes in B. pertussis, but a gradual decrease of virulence proteins. The quantity of virulence proteins concurred highly with the potency of the corresponding whole-cell pertussis vaccines, which were determined by the Kendrick test. In conclusion, proteome analysis provided detailed information on the composition and proportion of virulence proteins present in the whole-cell preparations of B. pertussis. Moreover, proteome analysis is a valuable method to monitor the production process of whole-cell biomass and predict the product quality of whole-cell pertussis vaccines.
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Bernard
authors.identifier: 0000-0001-6814-7656
authors.initials: B
authors.lastname: Metz
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Marieke
authors.initials: M
authors.lastname: Hoonakker
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Joost P
authors.initials: JP
authors.lastname: Uittenbogaard
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Michel
authors.initials: M
authors.lastname: Weyts
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Geert P M
authors.initials: GP
authors.lastname: Mommen
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Hugo D
authors.initials: HD
authors.lastname: Meiring
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Wichard
authors.initials: W
authors.lastname: Tilstra
authors.affiliation: National Institute for Public Health and the Environment , P.O. Box 1, 3720 BA Bilthoven, The Netherlands.
authors.forename: Jeroen L A
authors.initials: JL
authors.lastname: Pennings
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Leo A
authors.initials: LA
authors.lastname: van der Pol
authors.affiliation: National Institute for Public Health and the Environment , P.O. Box 1, 3720 BA Bilthoven, The Netherlands.
authors.forename: Betsy
authors.initials: B
authors.lastname: Kuipers
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Arjen
authors.initials: A
authors.lastname: Sloots
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Jan
authors.initials: J
authors.lastname: van den IJssel
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Bas
authors.initials: B
authors.lastname: van de Waterbeemd
authors.affiliation: Institute for Translational Vaccinology (Intravacc) , P.O. Box 450, 3720 AL Bilthoven, The Netherlands.
authors.forename: Arno
authors.initials: A
authors.lastname: van der Ark
mesh_terms: D000942:Antigens, Bacterial / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D061249:Batch Cell Culture Techniques; D019149:Bioreactors; D001886:Bordetella pertussis / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity; D002853:Chromatography, Liquid; D005285:Fermentation; D015870:Gene Expression; D006801:Humans; D008278:Magnesium Sulfate / Q000494:pharmacology; D013058:Mass Spectrometry; D037342:Pertussis Toxin / Q000037:antagonists & inhibitors / Q000096:biosynthesis* / Q000235:genetics; D010567:Pertussis Vaccine / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D020543:Proteome / Q000032:analysis* / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D014917:Whooping Cough / Q000276:immunology / Q000382:microbiology / Q000517:prevention & control
chemical_list: D000942:Antigens, Bacterial; D010567:Pertussis Vaccine; D020543:Proteome; D008278:Magnesium Sulfate; D037342:Pertussis Toxin
keywords: ELISA; LC−MS; continuous fermentation; microarray; potency; quality control; vaccine; whole-cell pertussis