Study reveals significant differences in protein expression between normal and colorectal cancer samples using two-dimensional difference gel electrophoresis
A new study published this month (March 2004) in Proteomics shows that two-dimensional difference gel electrophoresis (2D Dige) technology provided by Amersham Biosciences reveals statistically significant differences in protein abundance between normal and colorectal cancer samples that would not have been easily detected using classic 2D gel separation.
Ettan Dige technology from Amersham Biosciences has significant benefits over classic 2-D electrophoresis, providing a very sensitive and accurate method for measuring differences in protein expression. Researchers at Vanderbilt University medical centre, including members of the GI Spore, used 2D Dige coupled with mass spectrometry to investigate differences in protein expression between human colorectal cancers and normal mucosa.
Cy5-labelled proteins isolated from the tumour tissue of six patients and Cy3-labelled proteins isolated from adjacent normal mucosa were mixed together and then separated on the same 2D gel along with a Cy2-labelled mixture of all the samples.
Comparison with the mixed sample internal standard enabled protein abundance in the tumour and normal samples to be inter-compared among six patients without interference from gel-to gel variation.
Mass spectrometry and database interrogation were then used to identify those proteins that had a consistently different abundance in the tumour and normal samples.
Using this method, 52 unique proteins were identified that had a statistically significantly difference in abundance between tumour and normal samples.
For 42 of these 52 proteins, the use of Dige technology with a mixed sample internal standard enabled detection of protein abundance changes that would not have been evident if the Cy3-labelled and Cy5-labelled proteins from each patient had been compared separately.
"Identifying proteins that differ in abundance between normal and tumour samples may provide insight into critical events in cancer progression, or reveal proteins with potential value as therapeutic targets," said David Friedman, the study's lead investigator.
"Dige technology with a mixed sample internal standard enables us to find differences in protein abundance that would otherwise be overlooked due to the large degree of variation inherent between normal and tumour samples."
