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“Phagosome Closure Assay” to Visualize Phagosome Formation in Three Dimensions Using Total Internal Reflection Fluorescent Microscopy (TIRFM)
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Immunology and Infection
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JoVE Journal Immunology and Infection
“Phagosome Closure Assay” to Visualize Phagosome Formation in Three Dimensions Using Total Internal Reflection Fluorescent Microscopy (TIRFM)

“Phagosome Closure Assay” to Visualize Phagosome Formation in Three Dimensions Using Total Internal Reflection Fluorescent Microscopy (TIRFM)

DOI:
10.3791/54470-v

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10:07 min

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August 26, 2016

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, , ,
1Inserm U1016, Institut Cochin, CNRS UMR 8104,Université Paris Descartes, Sorbonne Paris Cité

Chapters

  • 00:05Title
  • 00:51Red Blood Cell (RBC) Opsonization and Poly-lysine Coverslip Coating
  • 02:31Non-covalent SRBC Fixation
  • 04:25TIRF Microscopy Phagocytosis Visualization
  • 05:23Image Acquisition and Processing
  • 07:45Results: F-actin Accumulation at the Site of Phagosome Closure
  • 08:51Conclusion

Summary

Automatic Translation

Automatic Translation

We describe an experimental setup to visualize with unprecedented high resolution phagosome formation and closure in three dimensions in living macrophages, using total internal reflection fluorescence microscopy. It allows monitoring of the base of the phagocytic cup, the extending pseudopods, as well as the precise site of phagosome scission.

Tags

Keywords: PhagosomeTIRFMTotal Internal Reflection Fluorescence Microscopy3D VisualizationMacrophagesPseudopod ExtensionTip FusionPlasma MembraneSheep Red Blood CellsSRBCsOpsonizationPolyL-lysineLive Cell Imaging

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