| Sensitive fluorescence detection of polyphosphate in polyacrylamide gels using 4',6-diamidino-2-phenylindol. | |
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MedLine Citation:
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PMID: 17847128 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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PAGE is commonly used to identify and resolve inorganic polyphosphates (polyP). We now report highly sensitive and specific staining methods for polyP in polyacrylamide gels based on the fluorescent dye, 4',6-diamidino-2-phenylindol (DAPI). DAPI bound to polyP in gels fluoresced yellow while DAPI bound to nucleic acids or glycosaminoglycans fluoresced blue. Inclusion of EDTA prevented staining of glycosaminoglycans by DAPI. We also identified conditions under which DAPI that was bound to polyP (but not nucleic acids or other anionic polymers) rapidly photobleached. This allowed us to develop an even more sensitive and specific negative staining method that distinguishes polyP from nucleic acids and glycosaminoglycans. The lower LOD using DAPI negative staining was 4 pmol (0.3 ng) phosphate per band, compared to conventional toluidine blue staining with a lower LOD of 250 pmol per band. |
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Authors:
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Stephanie A Smith; James H Morrissey |
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Publication Detail:
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Type: Journal Article; Research Support, N.I.H., Extramural; Research Support, Non-U.S. Gov't |
Journal Detail:
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Title: Electrophoresis Volume: 28 ISSN: 0173-0835 ISO Abbreviation: Electrophoresis Publication Date: 2007 Oct |
Date Detail:
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Created Date: 2007-10-08 Completed Date: 2008-02-06 Revised Date: 2010-08-24 |
Medline Journal Info:
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Nlm Unique ID: 8204476 Medline TA: Electrophoresis Country: Germany |
Other Details:
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Languages: eng Pagination: 3461-5 Citation Subset: IM |
Affiliation:
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Department of Internal Medicine, College of Medicine, University of Illinois at Urbana-Champaign, 506 S. Mathews, Urbana, IL 61801, USA. sasmith6@uiuc.edu |
Export Citation:
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APA/MLA Format Download EndNote Download BibTex |
| MeSH Terms | |
Descriptor/Qualifier:
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Edetic Acid
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chemistry Electrophoresis, Polyacrylamide Gel / methods* Fluorescent Dyes / chemistry* Glycosaminoglycans / chemistry Indoles / chemistry* Nucleic Acids / chemistry Polyphosphates / analysis*, chemistry* Sensitivity and Specificity Spectrometry, Fluorescence Staining and Labeling / methods* |
| Grant Support | |
ID/Acronym/Agency:
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HL 47014/HL/NHLBI NIH HHS; R01 HL047014-15/HL/NHLBI NIH HHS |
| Chemical | |
Reg. No./Substance:
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0/Fluorescent Dyes; 0/Glycosaminoglycans; 0/Indoles; 0/Nucleic Acids; 0/Polyphosphates; 47165-04-8/DAPI; 60-00-4/Edetic Acid |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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