Application of environmental scanning electron microscopy to determine biological surface structure

Area of Science:

• Cell Biology
• Microscopy Techniques
• Biophysics

Background:

• Environmental scanning electron microscopy (ESEM) is increasingly utilized in biological studies.
• Understanding the advantages and limitations of ESEM is crucial for its effective application.
• Previous research highlights specimen preparation artifacts in high-vacuum scanning electron microscopy (HV-SEM).

Purpose of the Study:

• To evaluate the impact of ESEM imaging on mammalian cell viability.
• To compare the challenges and artifacts associated with ESEM and HV-SEM.
• To explore the potential of ESEM for dynamic biological experiments.

Main Methods:

• Human monocyte-derived macrophages were used as a model system.
• Specimens were prepared for both ESEM and HV-SEM.
• Cell viability was assessed after ESEM imaging.
• Imaging of delicate cellular structures like membrane ruffles was performed.

Main Results:

• ESEM imaging was found to be detrimental to mammalian cell viability due to dehydration-induced damage.
• High-resolution imaging in ESEM presents unique challenges.
• Mammalian cells cannot be kept alive during ESEM imaging, likely due to membrane integrity loss during liquid removal.
• Fixation prior to imaging is recommended to mitigate damage.

Conclusions:

• ESEM and HV-SEM have complementary strengths and are best used in combination.
• Mammalian cells require fixation before ESEM imaging to prevent irreversible damage.
• ESEM is not suitable for live imaging of mammalian cells due to the dehydration process.