PXD000509

PXD000509 is an original dataset announced via ProteomeXchange.

Title	Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing
Description	Biotherapeutics are often produced in non-human host cells like Escherichia coli, yeast, and various mammalian cell lines. A major focus of any therapeutic protein purification process is to reduce host cell proteins to an acceptable low level. In this study, various E. coli host cell proteins were identified at different purifications steps by HPLC fractionation, SDS-PAGE analysis, and tryptic peptide mapping combined with online liquid chromatography mass spectrometry (LC-MS). However, no host cell proteins could be verified by direct LC-MS analysis of final drug substance material. In contrast, the application of affinity enrichment chromatography prior to comprehensive LC-MS was adequate to identify several low abundant host cell proteins at the final drug substance level. Bacterial alkaline phosphatase (BAP) was identified as being the most abundant host cell protein at several purification steps. Thus, we firstly established two different assays for enzymatic and immunological BAP monitoring using the cobas® technology. By using this strategy we were able to demonstrate an almost complete removal of BAP enzymatic activity by the established therapeutic protein purification process. In summary, the impact of fermentation, purification, and formulation conditions on host cell protein removal and biological activity can be conducted by monitoring process-specific host cell proteins in a GMP-compatible and high-throughput (> 1000 samples/day) manner.
HostingRepository	PRIDE
AnnounceDate	2014-07-24
AnnouncementXML	Submission_2014-07-24_03:57:01.xml
DigitalObjectIdentifier	https://dx.doi.org/10.6019/PXD000509
ReviewLevel	Peer-reviewed dataset
DatasetOrigin	Original dataset
RepositorySupport	Supported dataset by repository
PrimarySubmitter	Patrick Bulau
SpeciesList	scientific name: Escherichia coli; NCBI TaxID: 562;
ModificationList	No PTMs are included in the dataset
Instrument	LTQ Orbitrap; instrument model: LTQ Orbitrap

Revision	Datetime	Status	ChangeLog Entry
0	2013-10-09 07:41:03	ID requested
1	2014-04-15 04:13:56	announced
⏵ 2	2014-07-24 03:57:02	announced	Updated project metadata.

Bomans K, Lang A, Roedl V, Adolf L, Kyriosoglou K, Diepold K, Eberl G, M, ø, lh, ø, j M, Strauss U, Schmalz C, Vogel R, Reusch D, Wegele H, Wiedmann M, Bulau P, Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing. PLoS One, 8(11):e81639(2013) [pubmed]

curator keyword: Technical

submitter keyword: host cell protein

protein purification

mass spectrometry

affinity enrichment chromatography

cobas

high throughput immunochemistry testing

Patrick Bulau
contact affiliation	Roche
contact email	patrick.bulau@roche.com
dataset submitter

Dataset FTP location NOTE: Most web browsers have now discontinued native support for FTP access within the browser window. But you can usually install another FTP app (we recommend FileZilla) and configure your browser to launch the external application when you click on this FTP link. Or otherwise, launch an app that supports FTP (like FileZilla) and use this address: ftp://ftp.pride.ebi.ac.uk/pride/data/archive/2014/04/PXD000509

PRIDE project URI

• PRIDE
  1. PXD000509
     1. Label: PRIDE project
     2. Name: Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing