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An improved high throughput protein-protein interaction assay for nuclear hormone receptors
The Glutathione-S-Transferase (GST) “pulldown” assay has been used extensively to assay protein interactions in vitro. This methodology has been especially useful for investigating the interactions of nuclear hormone receptors with a wide variety of their interacting partners and coregulatory protei...
Autores principales: | , , |
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Formato: | Texto |
Lenguaje: | English |
Publicado: |
The Nuclear Receptor Signaling Atlas
2007
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1853068/ https://www.ncbi.nlm.nih.gov/pubmed/17464356 http://dx.doi.org/10.1621/nrs.05002 |
Sumario: | The Glutathione-S-Transferase (GST) “pulldown” assay has been used extensively to assay protein interactions in vitro. This methodology has been especially useful for investigating the interactions of nuclear hormone receptors with a wide variety of their interacting partners and coregulatory proteins. Unfortunately, the original GST-pulldown technique relies on multiple binding, washing and elution steps performed in individual microfuge tubes, and requires repeated centrifugation, aspiration, and suspension steps. This type of batch processing creates a significant liquid handling bottleneck, limiting the number of sample points that can be incorporated into one experiment and producing inherently less efficient washing and elution than would a flow-through methodology. In this manuscript, we describe the adaptation of this GST-pulldown assay to a 96-well filter plate format. The use of a multi-well filter plate makes it possible to assay more samples in significantly less time using less reagents and more efficient sample processing than does the traditional single tube assay. |
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