Change search
ReferencesLink to record
Permanent link

Direct link
Extracellular metabolite fingerprinting of spent mammalian cell culture medium in relation to the quality of an expressed recombinant protein
Stockholm University, Faculty of Science, Department of Analytical Chemistry.
(English)Manuscript (preprint) (Other academic)
Abstract [en]

This study demonstrates methodology for predicting the quality (in terms, for instance, of specific activities) of recombinant proteins expressed by mammalian cell cultures, at upstream process stages, using metabolic fingerprint analysis. Metabolites extracted from samples taken from a culture of a transfected Chinese Hamster Ovary (CHO) cell line expressing a recombinant protein in a bioreactor at various days of the cell culture process were analyzed by ultra high-pressure liquid chromatography-electrospray ionization-time of flight mass spectrometry (UHPLC-ESI-TOFMS). The LC-MS data were processed and the extracted information was correlated with the concentration of the active protein by partial least squares (PLS) regression, which revealed strong correlations between the LC-MS results and the concentration of active protein (R

2 = 0.99). The correlations between the LC-MS data and other parameters (glucose concentration, lactate concentration and the number of viable cells) were also studied. To obtain an overview of the data, Principal Component Analysis (PCA) was applied to the LC-MS data obtained from the samples to observe clustering or separation in the sample set. The PCA indicated that the LC-MS data obtained from samples from different days were significantly separated in temporal order from day 7 to day 28, according to the first Principal Component.

URN: urn:nbn:se:su:diva-28900OAI: diva2:227757
Available from: 2009-07-21 Created: 2009-07-16 Last updated: 2010-01-14Bibliographically approved
In thesis
1. Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
Open this publication in new window or tab >>Methods for structural studies of an antibody, screening metabolites in rat urine and analysis of spent cell cultivation media using LC/ESI-MS and chemometrics
2009 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes bioanalytical methods for generating fingerprints of biological systems for extracting relevant information with (protein) drugs in focus. Similarities and differences between samples can reveal the hidden relevant information, which can be used to optimize the production and facilitate the quality control of such protein drugs during their development and manufacture. Metabolic fingerprinting and multivariate data analysis (MVDA) can also facilitate early diagnosis of diseases and the effects and toxicity of drugs.

Currently, several protein drugs are available on the global market. Nevertheless, despite, the success of such biotherapeutics significant challenges remain to be overcome in maintaining their stability and efficacity throughout their production cycle and long-term storage. The native structure and functional activity of therapeutic proteins is affected by many variables from production to delivery, incl. variables assoc. with conditions in bioreactors, purification, storage and delivery. Thus, part of the work underlying this thesis focused on structural analysis of a protein drug using chemical labeling, peptide mapping, and evaluation of the charge state distributions of the whole protein generated by ESI. The other part focuses on non-targeted metabolomics with a view to optimizing the cell cultivation process and assessment of the drug’s toxicity. A combination of appropriate analytical methods and MVDA is needed to find markers that can facilitate optimization of the cultivation system and expression of the target proteins in early stages of process development. Rapid methods for characterizing the protein drugs in different stages of the process are also required for quality control.

In order to obtain high quality fingerprints analytical separation techniques with high resolution (such as HPLC or UHPLC) and sensitive analytical detection techniques (such as ESI, quadrupole or TOF MS) have been used, singly or in combination.

Place, publisher, year, edition, pages
Stockholm: Department of Analytical Chemistry, Stockholm University, 2009. 74 p.
Screening, Fingerprinting, metabolomics, Protein drugs, Mammalian Cell lines, LC/ESI-MS, chemometrics
National Category
Analytical Chemistry
Research subject
Analytical Chemistry
urn:nbn:se:su:diva-28921 (URN)978-91-7155-897-8 (ISBN)
Public defence
2009-09-16, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 12 A, Stockholm, 10:00 (English)
At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 3: Manuscript.Available from: 2009-08-25 Created: 2009-07-20 Last updated: 2009-08-27Bibliographically approved

Open Access in DiVA

No full text

Other links

Link to doctoral thesis

Search in DiVA

By author/editor
Zamani, Leila
By organisation
Department of Analytical Chemistry

Search outside of DiVA

GoogleGoogle Scholar
The number of downloads is the sum of all downloads of full texts. It may include eg previous versions that are now no longer available

Total: 74 hits
ReferencesLink to record
Permanent link

Direct link