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Related Concept Videos

Preparation of Samples for Electron Microscopy01:20

Preparation of Samples for Electron Microscopy

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To be visualized by an electron microscope, either transmission or scanning, biological samples need to be fixed (stabilized) so the electron beam does not destroy them and dried thoroughly (desiccated/dehydrated) so the vacuum does not affect them. Fixation needs to be done as quickly as possible because the sample properties will start changing as soon as it is removed from its natural environment. For example, in a tissue sample, the oxygen levels begin decreasing, causing an altered...
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Related Experiment Video

Updated: Jun 15, 2025

Imaging Replicative Domains in Ultrastructurally Preserved Chromatin by Electron Tomography
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A standardized protocol for sample preparation for scanning electron microscopy to visualize extrachromosomal DNA.

Jillann A Madren1, Jingting Chen2, William Dennis3

  • 1Microscopy Services Laboratory, University of North Carolina at Chapel Hill, Chapel Hill, NC 27516, USA.

Biotechniques
|August 26, 2024
PubMed
Summary

We developed a new scanning electron microscopy (SEM) protocol to visualize extrachromosomal DNA (ecDNA), including double minute (DM) chromosomes, in cancer research. This method enhances structural understanding of ecDNA, crucial for cancer and drug resistance studies.

Keywords:
correlative light and electron microscopydouble minute chromosomesextrachromosomal DNAfluorescence microscopyscanning electron microscopy

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Area of Science:

  • Molecular Biology
  • Genetics
  • Cancer Research

Background:

  • Extrachromosomal DNA (ecDNA) are circular DNA molecules frequently found in cancer cells.
  • Double minute (DM) chromosomes are a type of ecDNA linked to drug resistance and cancer progression.
  • Existing imaging techniques have limitations in preserving ecDNA structure during sample preparation.

Purpose of the Study:

  • To develop and optimize a systematic Scanning Electron Microscopy (SEM) protocol for high-resolution visualization of ecDNA.
  • To provide a standardized method for analyzing the circular architecture of ecDNA.
  • To improve the understanding of ecDNA's role in cancer and drug resistance.

Main Methods:

  • Development of a novel, end-to-end sample preparation protocol for SEM imaging.
  • Optimization of SEM parameters for visualizing fine ecDNA structures.
  • Application of the protocol to study ecDNA, including DM chromosomes.

Main Results:

  • Achieved high-resolution visualization of ecDNA structures using SEM.
  • Demonstrated the effectiveness of the optimized protocol in preserving ecDNA morphology.
  • Established a standardized method for ecDNA analysis.

Conclusions:

  • The developed SEM protocol offers a significant advancement in visualizing ecDNA.
  • This technique provides crucial insights into the structural characteristics of ecDNA.
  • The findings support further research into ecDNA's role in oncogenesis and therapeutic resistance.