Diffusion coefficients of proteins in carrier ampholyte versus immobiline gels

Cecilia Gelfi, Maria Luisa Bossi, Pier Giorgio Righetti

Research output: Contribution to journalArticle

Abstract

The apparent diffusion coefficients of proteins in carrier ampholyte isoelectric focusing (CA-IEF) and in immobilized pH gradients (IPGs) are strongly dependent on the amount of buffering ions present in the system. However, whereas in CA-IEF increased levels of ampholytes facilitate diffusion, in IPGs they strongly quench it. It is concluded that a protein in an IPG matrix is isoelectric but not isoionic, in the sense that it forms a salt with the surrounding ions bound to the polyacrylamide matrix. This salt formation is beneficial as it greatly increases protein solubility at the pI. It is suggested that, when performing zymograms in situ, the IPG gel should contain at least twice the standard amount of Immobiline, so as to keep sharp enzyme bands even with prolonged incubation periods.

Original languageEnglish
Pages (from-to)225-236
Number of pages12
JournalJournal of Chromatography A
Volume390
Issue number2
DOIs
Publication statusPublished - 1987

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ASJC Scopus subject areas

  • Analytical Chemistry

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