One-step split GFP staining for sensitive protein detection and localization in mammalian cells.
Biotechniques
; 49(4): 727-8, 730, 732 passim, 2010 Oct.
Article
em En
| MEDLINE
| ID: mdl-20964633
ABSTRACT
Although epitope tags are useful to detect intracellular proteins and follow their localization with antibodies, background and nonspecific staining often remain problematic. We describe a simple assay based on the split GFP complementation system. Proteins tagged with the 15-amino acid GFP 11 fragment are detected with a solution of the recombinant nonfluorescent complementary GFP 1-10 fragment to reconstitute a fluorescent GFP. In contrast to antibody-based staining methods, this one-step assay presents high specificity and very low background of fluorescence, thus conferring higher signal-to-noise ratios. We demonstrate that this new application of the split GFP tagging system facilitates detection of proteins displaying various subcellular localizations using flow cytometry and microscopy analysis.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Proteínas Recombinantes de Fusão
/
Espaço Intracelular
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Proteínas de Fluorescência Verde
Tipo de estudo:
Diagnostic_studies
Limite:
Animals
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Humans
Idioma:
En
Ano de publicação:
2010
Tipo de documento:
Article