Optimization of phosphopeptide elution conditions in immobilized Fe(III) affinity chromatography

A1 Originalartikel i en vetenskaplig tidskrift (referentgranskad)


Interna författare/redaktörer


Publikationens författare: Imanishi, Kochin, Eriksson
Publiceringsår: 2007
Tidskrift: Proteomics
Tidskriftsakronym: Proteomics
Volym: 7
Nummer: 2
Artikelns första sida, sidnummer: 174
Artikelns sista sida, sidnummer: 176
ISSN: 1615-9853
eISSN: 1615-9861


Abstrakt

While immobilized metal affinity chromatography (IMAC) has been widely used for affinity purification of phosphopeptides, the technique suffers from insufficient specificity. Therefore, there is an urgent need for IMAC optimization to yield the selectivity and sensitivity that is required for more challenging analyses. Recently, 2,5-dihydroxybenzoic acid (DHB) and phosphoric acid mixture has been reported as an efficient IMAC eluant. The disadvantage of DHB is that is not suitable for electrospray ionization-mass spectrometry. While further developing the IMAC elution protocol to overcome this problem, we noticed that DHB is not necessary and found a novel combination of phosphoric acid and acetonitrile to be more efficient. The purification efficacy of the novel protocol is superior to all previously described methods, while still being compatible with the most commonly used mass-spectrometric techniques in phosphoproteomics.

Senast uppdaterad 2019-19-09 vid 05:15