| Soft immobilized pH gradient gels in proteome analysis: a follow-up. | |
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MedLine Citation:
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PMID: 12833504 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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As a follow-up of a previous work on two-dimensional map analysis utilizing soft (< 4%T) immobilized pH gradient (IPG) matrices in the first dimension (Candiano et al., Electrophoresis 2002, 23, 292-297), we have further optimized the preparation of such dilute IPG gels. One important step for obtaining an even reswelling of the entire IPG strip along the pH 3-10 interval is a washing step in 100 mM citric acid. It appears as though after rinsing off the excess acid in distilled water, a gradient of this tricarboxylic acid remains trapped into the IPG matrix, from almost nil at the acidic gel region to substantially higher amounts in its basic counterpart. This gradient helps in obtaining a uniform reswelling of the IPG strip, since carboxyl groups are more heavily hydrated than amino groups. The combined effects of uniform reswelling and of diluting the gel matrix favor penetration of large macromolecules (> 200 kDa) and allow for better spot resolution and for the display of a substantially higher number of spots also in the 30-60 000 Da region. A delipidation step in tri-n-butylphosphate:acetone:methanol (1:12:1) appears to substantially improve spot focusing and greatly diminish streaking and smearing of spots in all regions of the pH gradient. |
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Authors:
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Maurizio Bruschi; Luca Musante; Giovanni Candiano; Gian Marco Ghiggeri; Ben Herbert; Francesca Antonucci; Pier Giorgio Righetti |
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Publication Detail:
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Type: Journal Article; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Proteomics Volume: 3 ISSN: 1615-9853 ISO Abbreviation: Proteomics Publication Date: 2003 Jun |
Date Detail:
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Created Date: 2003-06-30 Completed Date: 2004-03-24 Revised Date: 2006-11-15 |
Medline Journal Info:
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Nlm Unique ID: 101092707 Medline TA: Proteomics Country: Germany |
Other Details:
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Languages: eng Pagination: 821-5 Citation Subset: IM |
Affiliation:
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Laboratory on Physiopathology of Uremia, G. Gaslini Children's Hospital, Genoa, Italy. |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Blood Platelets
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chemistry* Gels / chemistry* Humans Hydrogen-Ion Concentration Molecular Weight Peptide Mapping Peptides / blood*, isolation & purification Proteome / analysis* Proton-Motive Force |
| Chemical | |
Reg. No./Substance:
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0/Gels; 0/Peptides; 0/Proteome |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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