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Nitrogen Cavitation and Differential Centrifugation Allows for Monitoring the Distribution of Peripheral Membrane Proteins in Cultured Cells
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JoVE Journal Biochimica
Nitrogen Cavitation and Differential Centrifugation Allows for Monitoring the Distribution of Peripheral Membrane Proteins in Cultured Cells

Nitrogen Cavitation and Differential Centrifugation Allows for Monitoring the Distribution of Peripheral Membrane Proteins in Cultured Cells

DOI:
10.3791/56037-v

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08:24 min

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August 18, 2017

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,
1Langone Medical Center,New York University

Capitoli

  • 00:05Titolo
  • 00:41Cell Harvesting
  • 01:35Nitrogen Cavitation
  • 03:31Separation of Cytosolic and Membrane Fractions
  • 05:22Results: Partitioning of Cellular Proteins in Membrane and Cytosolic Fractions
  • 07:07Conclusion

Summary

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Here we present protocols for detergent-free homogenization of cultured mammalian cells based on nitrogen cavitation and subsequent separation of cytosolic and membrane-bound proteins by ultracentrifugation. This method is ideal for monitoring the partitioning of peripheral membrane proteins between soluble and membrane fractions.

Tags

Nitrogen CavitationDifferential CentrifugationPeripheral Membrane ProteinsCultured CellsSubcellular CompartmentalizationHEK293 CellsHomogenization BufferPressurizationProtease InhibitorCell DisruptionFractionationSoluble FractionMembrane Fraction

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