Free flow-isoelectric focusing of human cellular lysates as sample preparation for protein analysis

✍ Scribed by Dorothe Burggraf; Gerhard Weber; Friedrich Lottspeich

• Publisher: John Wiley and Sons
• Year: 1995
• Tongue: English
• Weight: 671 KB
• Volume: 16
• Category: Article
• ISSN: 0173-0835
• DOI: 10.1002/elps.11501601169

✦ Synopsis

Abstract

A whole cell lysate of human cells was separated into 80 fractions according to the p__I__ of proteins using free flow isoelectric focusing with carrier ampholytes. The resolution of the process was highly reproducible, with an overlap of fractions of less than 30%. A protein of a faint silver stained spot in two‐dimensional gel electrophoresis (2‐DE) could be enriched, yielding a Coomassie blue stained spot which could be further characterized by proteinchemical methods. The enrichment of less abundant proteins from a complex crude cell extract was found to be a suitable tool for sample preparation and enrichment before applying proteins to 2‐DE and reversed‐phase high performance liquid chromatography.