The basis for heparin fractionation into 21 components by isoelectric focusing has been shown to be a strong interaction between the polysaccharide and different amphoteric species in the Ampholine mixture. This was demonstrated by altering the heparin/Ampholine ratio, by loading the sample either before focusing or to a prefocused gel slab and by re-running single heparin bands. The complexes exhibiting apparent pI values in the pH range 3.2--4.5 appear to be particularly stable, probably because an optimal amount of amino groups (4 to 5) in the Ampholine molecules are protonated. When stained with Toluidine blue, the heparin/Ampholine complexes precipitated in the gel exhibited different degrees of metachromasia, reflecting competition of the dye and individual components of Ampholine with respect to binding sites of heparin: at least three colours, violet, blue and indigo, are distinguishable. Ampholine, when added to a heparin . Toluidine blue complex in solution displaces the dye from the polysaccharide.

Isoelectric focusing of heparin. Evidence for complexing with carrier ampholytes / P.G. Righetti, E. Gianazza. - In: BIOCHIMICA ET BIOPHYSICA ACTA. PROTEIN STRUCTURE. - ISSN 0005-2795. - 532:1(1978 Jan 25), pp. 137-146.

Isoelectric focusing of heparin. Evidence for complexing with carrier ampholytes

E. Gianazza
Ultimo
1978

Abstract

The basis for heparin fractionation into 21 components by isoelectric focusing has been shown to be a strong interaction between the polysaccharide and different amphoteric species in the Ampholine mixture. This was demonstrated by altering the heparin/Ampholine ratio, by loading the sample either before focusing or to a prefocused gel slab and by re-running single heparin bands. The complexes exhibiting apparent pI values in the pH range 3.2--4.5 appear to be particularly stable, probably because an optimal amount of amino groups (4 to 5) in the Ampholine molecules are protonated. When stained with Toluidine blue, the heparin/Ampholine complexes precipitated in the gel exhibited different degrees of metachromasia, reflecting competition of the dye and individual components of Ampholine with respect to binding sites of heparin: at least three colours, violet, blue and indigo, are distinguishable. Ampholine, when added to a heparin . Toluidine blue complex in solution displaces the dye from the polysaccharide.
Swine ; Animals ; Kinetics ; Binding, Competitive ; Spectrophotometry ; Tolonium Chloride ; Isoelectric Focusing ; Heparin
Settore BIO/10 - Biochimica
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2434/176209
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