Protein extraction and 2-DE of water- and lipid-soluble proteins from bovine pericardium, a low-cellularity tissue

Leigh G. Griffiths, Leila Choe, Kelvin H. Lee, Kenneth F. Reardon, E. Christopher Orton

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

Bovine pericardium (BP) is an important biomaterial used in the production of glutaraldehyde-fixed heart valves and tissue-engineering applications. The ability to perform proteomic analysis on BP is useful for a range of studies, including investigation of immune rejection after implantation. However, proteomic analysis of fibrous tissues such as BP is challenging due to their relative low-cellularity and abundance of extracellular matrix. A variety of methods for tissue treatment, protein extraction, and; fractionation were investigated with the aim of producing high-quality 2-DE gels for both water- and lipid-soluble BP proteins. Extraction of water-soluble proteins with 3-(benzyldimethylammonio)-propanesulfonate followed by n-dodecyl β-D-maltoside extraction and ethanol precipitation for lipid-soluble proteins provided the best combination of yield, spot number, and resolution on 2-DE gels (Protocol E2). ESI-quadrupole/ion trap or MALDI-TOF/TOF MS protein identifications were performed to confirm bovine origin and appropriate subcellular prefractionation of resolved proteins. Twenty-five unique, predominantly cytoplasmic bovine proteins were identified from the water-soluble fraction. Thirty-two unique, predominantly membrane bovine proteins were identified from the lipid-soluble fraction. These results demonstrated that the final protocol produced high-quality proteomic data from this important tissue for both cytoplasmic and membrane proteins.

Original languageEnglish (US)
Pages (from-to)4508-4515
Number of pages8
JournalElectrophoresis
Volume29
Issue number22
DOIs
StatePublished - Dec 1 2008

Keywords

  • 2-DE
  • Biological scaffold
  • Bovine pericardium
  • Protein extraction

ASJC Scopus subject areas

  • Analytical Chemistry
  • Biochemistry
  • Clinical Biochemistry

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