IEF is often used in multidimensional shotgun proteomics and the narrow range of 3.5-4.5 is the recommended pH interval for the fractionation of tryptic peptides. Usually, even if IEF is performed in IPG strip with a narrow range pH, the entire sample must be loaded onto the strip, including the "out of IPG range" peptides. We describe a simple protocol to recover at least a part of these missing peptides and show that this recovery significantly influences the overall fractionation result, increasing the number of the identified proteins and the protein coverage.
No peptide left behind: the "out of range" recovery in IPG-IEF fractionation.
IORI, ELISABETTA;RATTAZZI, MARCELLO;MILLIONI, RENATO
2014
Abstract
IEF is often used in multidimensional shotgun proteomics and the narrow range of 3.5-4.5 is the recommended pH interval for the fractionation of tryptic peptides. Usually, even if IEF is performed in IPG strip with a narrow range pH, the entire sample must be loaded onto the strip, including the "out of IPG range" peptides. We describe a simple protocol to recover at least a part of these missing peptides and show that this recovery significantly influences the overall fractionation result, increasing the number of the identified proteins and the protein coverage.File in questo prodotto:
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