Epithelial cell preparation for proteomic and transcriptomic analysis in human pancreatic tissue

Pathology, Research and Practice
Udo KellnerM A Reymond

Abstract

Standardized sample preparation procedures constitute a prerequisite for obtaining reliable and reproducible results in gene expression research in humans. In particular, in diseases such as pancreatic cancer and pancreatitis, isolating epithelial cells is an important step preceding such research. In pancreatic tissue, the high amount of RNAases is a further problem when it comes to obtaining high-quality RNA, and the presence of secreted proteases accelerates protein degradation. We developed a successful method that addresses these different problems. This method, which uses epithelial cell surface antibody Ber-Ep4, proteases, and RNAases inhibitors, leads to a significant enrichment (> 95% purity) of epithelial cells from fresh human tissue samples and allows for both proteomics (Western Blot, 2D PAGE) and transcriptomics studies (rtPCR, cDNA microarray). Compared with other cell purification procedures, this method is characterized by several advantages: a large quantity of cells available for downstream analysis, combined transcriptomics and proteomics studies using the same samples, better reproducibility of proteomics studies, and an acceptable yield (63%) for gene expression arrays studies. Moreover, a quality control ...Continue Reading

References

Aug 5, 2000·Virchows Archiv : an International Journal of Pathology·J PetersG Klöppel
Oct 10, 2003·American Journal of Physiology. Cell Physiology·Tamás OrdögKenton M Sanders

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Citations

Apr 2, 2010·Tumour Biology : the Journal of the International Society for Oncodevelopmental Biology and Medicine·Marta Sanchez-Carbayo
Mar 1, 2008·Expert Opinion on Medical Diagnostics·Marta Sanchez-Carbayo
Jul 9, 2004·Pathology, Research and Practice·Christoph RöckenAlbert Roessner
Feb 3, 2005·Molecular & Cellular Proteomics : MCP·Ru ChenRuedi Aebersold
Mar 25, 2005·Proteomics·Matthias P A EbertChristoph Röcken
Jul 26, 2005·Proteomics·Emil SagynalievMarc-André Reymond

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