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Micropatterning Transmission Electron Microscopy Grids to Direct Cell Positioning within Whole-Cell Cryo-Electron Tomography Workflows
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Biochemistry
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JoVE Journal Biochemistry
Micropatterning Transmission Electron Microscopy Grids to Direct Cell Positioning within Whole-Cell Cryo-Electron Tomography Workflows

Micropatterning Transmission Electron Microscopy Grids to Direct Cell Positioning within Whole-Cell Cryo-Electron Tomography Workflows

DOI:
10.3791/62992-v

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09:53 min

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September 13, 2021

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, , ,
1Department of Biochemistry,University of Wisconsin, Madison, 2Cryo-Electron Microscopy Research Center, Department of Biochemistry,University of Wisconsin, Madison, 3Midwest Center for Cryo-Electron Tomography, Department of Biochemistry,University of Wisconsin, Madison, 4Department of Chemistry,University of Wisconsin, Madison, 5Morgridge Institute for Research

Chapters

  • 00:04Introduction
  • 01:25Preparation of Grids for Patterning and Application of the Anti-fouling Layer
  • 02:50Applying PLPP Gel
  • 03:33Micropatterning
  • 06:11Deposition of Extracellular Matrix (ECM) Proteins
  • 06:54Results: Micropatterning to Direct Cell Positioning within Whole-cell Cryo-electron Tomography
  • 09:17Conclusion

Summary

Automatic Translation

Automatic Translation

The goal of this protocol is to direct cell adhesion and growth to targeted areas of grids for cryo-electron microscopy. This is achieved by applying an anti-fouling layer that is ablated in user-specified patterns followed by deposition of extra-cellular matrix proteins in the patterned areas prior to cell seeding.

Tags

Keywords: MicropatterningTransmission Electron MicroscopyCryo-electron TomographyCell PositioningWhole-cell Cryo-electron TomographyEM GridsPoly-L-lysinePEG-SVAHEPESCell CultureCryo-CLEMCryo-FIB Milling

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