FIB-SEM as a Volume Electron Microscopy Approach to Study Cellular Architectures in SARS-CoV-2 and Other Viral Infections: A Practical Primer for a Virologist
Valentina Baena,
Ryan Conrad,
Patrick Friday,
Ella Fitzgerald,
Taeeun Kim,
John Bernbaum,
Heather Berensmann,
Adam Harned,
Kunio Nagashima,
Kedar Narayan
Affiliations
Valentina Baena
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Ryan Conrad
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Patrick Friday
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Ella Fitzgerald
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Taeeun Kim
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
John Bernbaum
National Institute of Allergy and Infectious Diseases, Division of Clinical Research, Integrated Research Facility at Fort Detrick (IRF-Frederick), Frederick, MD 21702, USA
Heather Berensmann
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Adam Harned
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Kunio Nagashima
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
Kedar Narayan
Center for Molecular Microscopy, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA
The visualization of cellular ultrastructure over a wide range of volumes is becoming possible by increasingly powerful techniques grouped under the rubric “volume electron microscopy” or volume EM (vEM). Focused ion beam scanning electron microscopy (FIB-SEM) occupies a “Goldilocks zone” in vEM: iterative and automated cycles of milling and imaging allow the interrogation of microns-thick specimens in 3-D at resolutions of tens of nanometers or less. This bestows on FIB-SEM the unique ability to aid the accurate and precise study of architectures of virus-cell interactions. Here we give the virologist or cell biologist a primer on FIB-SEM imaging in the context of vEM and discuss practical aspects of a room temperature FIB-SEM experiment. In an in vitro study of SARS-CoV-2 infection, we show that accurate quantitation of viral densities and surface curvatures enabled by FIB-SEM imaging reveals SARS-CoV-2 viruses preferentially located at areas of plasma membrane that have positive mean curvatures.
