PXD000509 is an
original dataset announced via ProteomeXchange.
Dataset Summary
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 |
Dataset History
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. |
Publication List
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] |
Keyword List
curator keyword: Technical |
submitter keyword: host cell protein |
protein purification |
mass spectrometry |
affinity enrichment chromatography |
cobas |
high throughput immunochemistry testing |
Contact List
Patrick Bulau |
contact affiliation | Roche |
contact email | patrick.bulau@roche.com |
dataset submitter | |
Full Dataset Link List
Dataset FTP location
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PRIDE project URI |
Repository Record List
[ + ]
[ - ]
- PRIDE
- PXD000509
- Label: PRIDE project
- Name: Identification and monitoring of host cell proteins by mass spectrometry combined with high performance immunochemistry testing