Short monolithic columns for purification and fractionation of peptide samples for matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry analysis in proteomics
Jazyk angličtina Země Nizozemsko Médium print-electronic
Typ dokumentu hodnotící studie, časopisecké články, práce podpořená grantem
PubMed
19217112
DOI
10.1016/j.chroma.2009.01.075
PII: S0021-9673(09)00137-X
Knihovny.cz E-zdroje
- MeSH
- chromatografie kapalinová přístrojové vybavení metody MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- ječmen (rod) chemie MeSH
- jedlá semena chemie MeSH
- methakryláty chemie MeSH
- oxid křemičitý chemie MeSH
- peptidy analýza izolace a purifikace MeSH
- proteomika metody MeSH
- reprodukovatelnost výsledků MeSH
- sérový albumin hovězí analýza izolace a purifikace MeSH
- skot MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice metody MeSH
- zvířata MeSH
- Check Tag
- skot MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- hodnotící studie MeSH
- práce podpořená grantem MeSH
- Názvy látek
- dodecyl methacrylate MeSH Prohlížeč
- ethylene dimethacrylate MeSH Prohlížeč
- methakryláty MeSH
- oxid křemičitý MeSH
- peptidy MeSH
- sérový albumin hovězí MeSH
This study records a novel application of methacrylate-based monolithic columns for MALDI-TOF/TOF MS analyses in proteomics for pre-concentration and separation of peptides derived from protein digestion. Reversed-phase monolithic capillary columns (30mm x 0.32mm i.d.) were created inside the fused silica capillary via thermal-initiated free-radical polymerization of ethylene glycol dimethacrylate and lauryl methacrylate monomers in the presence of 1-propanol and 1,4-butandiol as a porogen system. The elution of peptides was achieved using a linear gradient of acetonitrile from 0 to 60% in water with 0.1% trifluoroacetic acid formed in a microsyringe. Individual fractions of separated peptides were collected on the MALDI target spots covered with alpha-cyano-4-hydroxycinnamic acid used as a matrix and then they were analyzed using MALDI-TOF/TOF mass spectrometry. The developed method was tested with a mixture of tryptic peptides from bovine serum albumin and its applicability was also tested for tryptic in-gel digests from barley grain extracts of water soluble proteins separated using SDS gel electrophoresis. The number of detected peptides was approximately three to four times higher compared to the analysis without previous separation. These results show an improved quality of sample information with the higher amount of identified peptides which increased protein sequence coverage and improved sensitivity of mass spectrometry measurements.
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