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J Biomed Sci. 2010 May 11;17:36. doi: 10.1186/1423-0127-17-36.

Trypsin-induced proteome alteration during cell subculture in mammalian cells.

Author information

1
Institute of Bioinformatics and Structural Biology & Department of Life Sciences, National Tsing Hua University, Hsinchu, Taiwan.

Abstract

BACKGROUND:

It is essential to subculture the cells once cultured cells reach confluence. For this, trypsin is frequently applied to dissociate adhesive cells from the substratum. However, due to the proteolytic activity of trypsin, cell surface proteins are often cleaved, which leads to dysregulation of the cell functions.

METHODS:

In this study, a triplicate 2D-DIGE strategy has been performed to monitor trypsin-induced proteome alterations. The differentially expressed spots were identified by MALDI-TOF MS and validated by immunoblotting.

RESULTS:

36 proteins are found to be differentially expressed in cells treated with trypsin, and proteins that are known to regulate cell metabolism, growth regulation, mitochondrial electron transportation and cell adhesion are down-regulated and proteins that regulate cell apoptosis are up-regulated after trypsin treatment. Further study shows that bcl-2 is down-regulated, p53 and p21 are both up-regulated after trypsinization.

CONCLUSIONS:

In summary, this is the first report that uses the proteomic approach to thoroughly study trypsin-induced cell physiological changes and provides researchers in carrying out their experimental design.

PMID:
20459778
PMCID:
PMC2873939
DOI:
10.1186/1423-0127-17-36
[Indexed for MEDLINE]
Free PMC Article
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