Article ID | Journal | Published Year | Pages | File Type |
---|---|---|---|---|
6491530 | Journal of Biotechnology | 2014 | 7 Pages |
Abstract
In this article, we describe and evaluate a two-step optimized protocol based on fast filtration by use of a vacuum pump for quenching and subsequent extraction of intracellular metabolites from CHO (Chinese hamster ovary) suspension cells, which uses commercially available components. The method allows transfer of washed cells into liquid nitrogen within 10-15Â s of sampling and recovers the entire extraction solution volume. It also has the advantage to remove residual filter filaments in the final sample, thus preventing damage to separation columns during subsequent MS analysis. Relative to other methods currently used in the literature, the resulting energy charge of intracellular adenosine nucleotides was increased to 0.94 compared to 0.90 with cold PBS quenching or 0.82 with cold methanol/AMBIC quenching.
Keywords
Related Topics
Physical Sciences and Engineering
Chemical Engineering
Bioengineering
Authors
Juan A. Hernández Bort, Vinoth Shanmukam, Martin Pabst, Markus Windwarder, Laura Neumann, Ali Alchalabi, Guido Krebiehl, Gunda Koellensperger, Stephan Hann, Denise Sonntag, Friedrich Altmann, Christine Heel, Nicole Borth,