Abstract
Characterisation of tyrosine phosphorylations induced in immune cells in response to inflammatory stimuli may help elucidate the molecular bases of the diversity of immune responses. We have used anti-phosphotyrosine antibodies in combination with cell surface biotinylation in a two-step affinity purification procedure to recover pervanadate-induced tyrosine phosphorylated proteins from sub-cellular compartments, including the cell surface, of murine T cells and macrophages prior to separation by solution-phase isoelectric focussing and one-dimensional gel electrophoresis and identification by tandem mass spectrometry.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Animals
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Antibodies
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Biotinylation
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Cell Compartmentation
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Cell Line
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Electrophoresis, Polyacrylamide Gel
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Hybridomas / metabolism
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Immunoprecipitation
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Isoelectric Focusing
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Macrophages / metabolism
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Mass Spectrometry
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Membrane Proteins / isolation & purification*
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Membrane Proteins / metabolism
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Mice
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Phosphoproteins / isolation & purification*
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Phosphoproteins / metabolism
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Phosphorylation
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Phosphotyrosine / immunology
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Phosphotyrosine / metabolism
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Protein Tyrosine Phosphatases / antagonists & inhibitors
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Proteome / metabolism*
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T-Lymphocytes / metabolism
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Vanadates / pharmacology
Substances
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Antibodies
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Membrane Proteins
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Phosphoproteins
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Proteome
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pervanadate
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Phosphotyrosine
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Vanadates
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Protein Tyrosine Phosphatases