The use of titanium dioxide micro-columns to selectively isolate phosphopeptides from proteolytic digests.
Methods Mol Biol
; 527: 57-66, xi, 2009.
Article
en En
| MEDLINE
| ID: mdl-19241005
ABSTRACT
Titanium dioxide has very high affinity for phosphopeptides and it has become an efficient alternative to already existing methods for phosphopeptide enrichment from complex samples. Peptide loading in a highly acidic environment in the presence of 2,5-dihydroxybenzoic acid (DHB), phthalic acid, or glycolic acid has been shown to improve selectivity significantly by reducing unspecific binding from nonphosphorylated peptides. The enriched phosphopeptides bound to the titanium dioxide are subsequently eluted from the micro-column using an alkaline buffer. Titanium dioxide chromatography is extremely tolerant towards most buffers used in biological experiments. It is highly robust and as such it has become one of the methods of choice in large-scale phospho-proteomics. Here we describe the protocol for phosphopeptide enrichment using titanium dioxide chromatography followed by desalting and concentration of the sample by reversed phase chromatography prior to MS analysis.
Texto completo:
1
Banco de datos:
MEDLINE
Asunto principal:
Péptido Hidrolasas
/
Fosfopéptidos
/
Titanio
/
Cromatografía en Gel
Límite:
Animals
/
Humans
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2009
Tipo del documento:
Article
País de afiliación:
Dinamarca