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Updated: Jun 22, 2026
Correlative Microscopy for 3D Structural Analysis of Dynamic Interactions
Published on: June 24, 2013
A polarized cell system amenable to subcellular resolution imaging of influenza virus infection
Jean-Baptiste Brault1, Catherine Thouvenot2, Magda Cannata Serio1
1Institut Pasteur, Université Paris Cité, CNRS UMR 3569, RNA Biology of Influenza Viruses, Paris, France.
Insights
This study introduces a 3D cell culture model for influenza A virus (IAV) infection, enabling better visualization of viral genome trafficking within polarized respiratory cells. This advanced model overcomes limitations of traditional 2D cultures for studying IAV pathogenesis.
Area of Science:
- Virology
- Cell Biology
- Biotechnology
Background:
- Influenza A virus (IAV) intracellular trafficking relies on host cell cytoskeleton and endomembrane systems.
- Conventional 2D cell cultures lack the polarized architecture of natural respiratory epithelial cells, limiting mechanistic studies of IAV infection.
- Subcellular imaging of IAV in 2D models does not reflect in vivo intracellular organization.
Purpose of the Study:
- To develop a 3D cell culture system for studying influenza A virus infection.
- To enable subcellular imaging along the baso-lateral axis of polarized cells during IAV infection.
- To provide a more physiologically relevant model for IAV pathogenesis research.
Main Methods:
- Culturing polarized Caco2-TC7 epithelial cell monolayers on Cytodex-3 microcarrier beads.
- Infecting these 3D-cultured cells with influenza A virus.
- Performing immunostaining, confocal imaging, and electron microscopy on infected polarized cells.
Main Results:
- Successful establishment of a 3D cell culture system for polarized epithelial cells infected with IAV.
- Demonstration of subcellular resolution imaging along the baso-lateral axis in the 3D model.
- Validation of the protocol for visualizing IAV-host cell interactions in a polarized context.
Conclusions:
- The developed 3D cell culture assay overcomes limitations of 2D models for studying IAV intracellular trafficking.
- This method allows for advanced imaging of IAV-infected polarized human epithelial cells.
- The protocol is adaptable for studying other pathogens that infect polarized epithelial cells.
Abstract:
The life cycle of influenza A viruses (IAV), and notably intracellular trafficking of the viral genome, depends on multiple interactions with the cellular cytoskeleton and endomembrane system. A limitation of the conventional cellular models used for mechanistic study and subcellular imaging of IAV infection is that they are cultured in two dimensions (2D) under non-polarizing conditions, and therefore they do not recapitulate the intracellular organization of the polarized respiratory epithelial cells naturally targeted by IAVs. To overcome this limitation, we developed an IAV-infection assay in a 3D cell culture system which allows imaging along the baso-lateral axis of polarized cells, with subcellular resolution. Here we describe a protocol to grow polarized monolayers of Caco2-TC7 cells on static Cytodex-3 microcarrier beads, infect them with IAV, and subsequently perform immunostaining and confocal imaging, or electron microscopy, on polarized IAV-infected cells. This method can be extended to other pathogens that infect human polarized epithelial cells.
