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A Western Blotting Protocol for Small Numbers of Hematopoietic Stem Cells
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Biochemistry
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JoVE Journal Biochemistry
A Western Blotting Protocol for Small Numbers of Hematopoietic Stem Cells

A Western Blotting Protocol for Small Numbers of Hematopoietic Stem Cells

DOI:
10.3791/56855-v

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08:45 min

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August 22, 2018

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, ,
1Division of Experimental Hematology,Cincinnati Children’s Hospital Medical Center, 2Department of Cell and Developmental Biology, Abramson Family Cancer Research Institute,Perelman School of Medicine, University of Pennsylvania, 3Department of Obstetrics and Gynecology, Southwest Hospital,Third Military Medical University

Chapters

  • 00:04Title
  • 00:48Sample Preparation
  • 04:20Electrophoresis, Transfer, and Block
  • 05:30Antibody Labeling and Detection
  • 06:09Results: Use of Optimized Western Blotting Technique to Analyze Small Numbers of Hematopoietic Stem and Progenitor Cells
  • 07:56Conclusion

Summary

Automatic Translation

Automatic Translation

A standard Western blotting protocol was optimized for analyzing as few as 500 hematopoietic stem or progenitor cells. Optimization involves careful handling of the cell sample, limiting transfers between tubes, and directly lysing the cells in Laemmli sample buffer.

Tags

Keywords: Western BlottingHematopoietic Stem CellsProtein AnalysisCell SortingProtein QuantificationLow Cell NumberCell LysisSample PreparationCentrifugationPhosphatase InhibitorsProtease InhibitorsLaemmli Buffer

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