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Chapters
- 00:00Title
- 00:41Introduction
- 01:13Preparing for Cross-linking
- 03:08Cross-linking and Cell Lysis
- 05:59Immunoprecipitation and Elution of the Bound Proteins
- 09:28IP of Cross-linked Complexes Representative Results
- 10:14Conclusion
The cell permeable crosslinker DSP [dithiobis-(succinimidyl propionate)] stabilizes transient and labile interactions in vivo, which allows their isolation using stringent protein complex purification techniques. Here we present a technique for crosslinking cells grown in culture followed by isolation of protein complexes by immunoprecipitation.
Tags
IsolationLabile Multi-protein ComplexesIn VivoControlled Cellular Cross-linkingImmuno-magnetic Affinity ChromatographyDynamic NatureTransient Protein AssociationsWeak Protein AssociationsLow Affinity InteractionsIsolation MethodsIdentification MethodsProtein NetworksChemical CrosslinkersSelective StabilizationBiochemical LimitationsPurification MethodsProtocolCells In CultureHomobifunctional CrosslinkerSpacer ArmDithiobis-(succinimidyl Proprionate)CleavageReductionDisulphide BondCross-linking And Immunoaffinity ChromatographyMagnetic BeadsProtein ComplexesPurification TechniquesWestern Blot TechniquesMass Spectrometry
