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Researchers have provided the first online searchable high-resolution 2D proteome of the human colon crypt to provide a tool for other proteomics researchers looking for biomarkers and drug targets.
The latest study, published in an advance article in the Journal of Proteome Research, indicates the relative order of protein abundance in human colon crypts obtained 10 to 20 cm away from the site of tumours from patients undergoing surgical resection for cancer of the rectum or sigmoid colon.
Colon cancer is the third most common type of cancer with nearly a million cases of colon cancer reported a year worldwide.
There are drugs on the market for sufferers; however patient prognosis is much better if the condition is diagnosed early - making the quest for biomarkers that could be used in a diagnostic setting particularly important.
The US researchers, led by Dr Anthony Yeung of the Fox Chase Cancer Center, believe that as the proteome coverage reaches thousands of proteins, it is essential to output the proteome maps and gene ontology in a searchable manner that can be hyperlinked to external databases - either one protein at a time or as a group of proteins.
As such they have provided the data on their website to help in the search for new cancer targets and biomarkers.
The researchers feel that several previous proteomic studies of the colon have been handicapped by the inclusion of both epithelial and mesenchymal cell types and an abbreviated pH range leading to the identification of a limited number of proteins and isoforms.
They therefore ensured that their study included only epithelial cells derived from colonic crypts and used two-dimensional gel electrophoresis (2D GE) at three overlapping pH ranges (pH 4-11) to ensure as complete protein coverage as possible.
Protein samples were then identified and excised from the gels before in-gel tryptic digestion.
The digested proteins were analysed by MALDI-TOF (matrix assisted laser desorption ionisation - time of flight) mass spectrometry (MS) to yield the peptide mass fingerprints of 1695 proteins.
This allowed the identification of approximately 800 nonredundant proteins and 900 isoforms from the human colonic crypts
The researches noted that the resolution of thousands of proteins by 2D GE does have its shortcomings, with the possibility that overlapping gels of three pH ranges could inflate the number of isoforms identified for a given protein.
They also mention that any given spot may overlap with one or more proteins, with MS analysis indicating that 18 per cent of the spots contained two proteins with the more prominent protein potentially masking the protein of interest.
They suggest this problem could be overcome by using gels of narrower pH ranges or using Western blotting.
