Skip to main content
padlock icon - secure page this page is secure

Monitoring of Glycoprotein Products in Cell Culture Lysates Using Lectin Affinity Chromatography and Capillary HPLC Coupled to Electrospray Linear Ion Trap‐Fourier Transform Mass Spectrometry (LTQ/FTMS)

Buy Article:

$59.00 + tax (Refund Policy)

In this paper, we describe the combination of lectin chromatography with capillary LC coupled to a linear ion trap‐Fourier transform mass spectrometer (LTQ/FTMS) to enrich and characterize overexpressed glycoproteins from a cell culture lysate. A well‐characterized glycoprotein, recombinant tissue plasminogen activator (rt‐PA), was used as a standard, and we demonstrated that the three N‐linked glycopeptides (including glycan structures) present in a tryptic digest of the rt‐PA standard could be characterized in the new hybrid MS platform. A feature of this approach is that a significant amount of information can be obtained about the carbohydrate structures by direct analysis of the tryptic digest without the need for additional time‐consuming sample preparation protocols. A combination of lectins was then studied for improved recovery of captured glycopeptides and was related to the selectivity of different lectins for specific glycosylation motifs. This approach was then extended to the lysate of a cell line routinely used in biotechnology manufacture (Chinese hamster ovary, CHO). This study showed that the combinations of lectins could enrich glycoproteins significantly from a CHO cell lysate. We also demonstrated that with this level of enrichment and with the new hybrid mass spectrometer, we could study the structures of N‐linked glycopeptides of rt‐PA present in a crude CHO cell lysate, at a ratio of 1:200 (rtPA:total cell lysate protein, w/w) by accurate mass measurement in the FTMS and tandem MS n in the linear ion trap. The generic and high throughput nature of the lectin approach combined with the ability to directly analyze the glycan structures in the tryptic digest suggest that this platform has the potential to routinely monitor glycoprotein products at early stage manufacturing in the biotech industry.
No References
No Citations
No Supplementary Data
No Article Media
No Metrics

Document Type: Research Article

Affiliations: Barnett Institute, Northeastern University, 360 Huntington Ave., Boston, Massachusetts 02115

Publication date: January 1, 2006

  • Access Key
  • Free content
  • Partial Free content
  • New content
  • Open access content
  • Partial Open access content
  • Subscribed content
  • Partial Subscribed content
  • Free trial content
Cookie Policy
X
Cookie Policy
Ingenta Connect website makes use of cookies so as to keep track of data that you have filled in. I am Happy with this Find out more