A new method for peptide analysis and purification is described, based on isoelectric focusing in immobilized pH gradients. On the analytical scale, the peptide zones can now be revealed by an stain for primary and secondary amino group (e.g. ninydrin, fluorescamine, dansyl chloride) since the buffering species, unlike conventional carrier ampholytes, contain only carboxyl and tertiary amino groups. For preparative purposes, conditions have been described to remove most contaminants (e.g. unreacted monomers, non-cross-linked, short polyacrylamide chains) from the gel matrix before the electrophoretic run. However, ca. 2% of the gel dry mass is still present as extractable material. The focused peptides can be recovered in higly yields (ca. 90%) with a fairly high degree of purity (75%), the contaminants being mostly components eluted from the polyacrylamide gel.

Analytical and preparative isoelectric focusing of peptides in immobilized pH gradients / E. Gianazza, F. Chillemi, M. Duranti, P.G. Righetti. - In: JOURNAL OF BIOCHEMICAL AND BIOPHYSICAL METHODS. - ISSN 0165-022X. - 8:4(1983), pp. 339-351.

Analytical and preparative isoelectric focusing of peptides in immobilized pH gradients

E. Gianazza;M. Duranti;
1983

Abstract

A new method for peptide analysis and purification is described, based on isoelectric focusing in immobilized pH gradients. On the analytical scale, the peptide zones can now be revealed by an stain for primary and secondary amino group (e.g. ninydrin, fluorescamine, dansyl chloride) since the buffering species, unlike conventional carrier ampholytes, contain only carboxyl and tertiary amino groups. For preparative purposes, conditions have been described to remove most contaminants (e.g. unreacted monomers, non-cross-linked, short polyacrylamide chains) from the gel matrix before the electrophoretic run. However, ca. 2% of the gel dry mass is still present as extractable material. The focused peptides can be recovered in higly yields (ca. 90%) with a fairly high degree of purity (75%), the contaminants being mostly components eluted from the polyacrylamide gel.
immobilized pH gradient; isoelectic focusing; peptide
Settore BIO/10 - Biochimica
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Utilizza questo identificativo per citare o creare un link a questo documento: http://hdl.handle.net/2434/177474
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