Light-sheet Microscopy for Three-dimensional Visualization of Human Immune Cells

Rouven Schoppmeyer; Renping Zhao; Markus Hoth; Bin Qu

doi:10.3791/57651

Microscopia de luz-folha para visualização tridimensional de células do sistema imunológico humanas

JoVE Journal
Immunology and Infection

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JoVE Journal Immunology and Infection

Light-sheet Microscopy for Three-dimensional Visualization of Human Immune Cells

Microscopia de luz-folha para visualização tridimensional de células do sistema imunológico humanas

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09:44 min

June 13, 2018

DOI: 10.3791/57651-v

Rouven Schoppmeyer¹, Renping Zhao¹, Markus Hoth¹, Bin Qu¹

¹Department of Biophysics, Center for Integrative Physiology and Molecular Medicine (CIPMM), School of Medicine,Saarland University

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Overview

This article presents a protocol for visualizing immune cells within a three-dimensional collagen matrix using light-sheet microscopy. It also details methods for tracking cell migration in this 3D environment.

Key Study Components

Area of Science

Immunology
Cell Biology
Microscopy Techniques

Background

Understanding immune cell behavior in 3D environments is crucial for immunology.
Light-sheet microscopy allows for selective illumination of cells.
This method minimizes photobleaching and cytotoxicity.
Tracking cell migration can provide insights into immune responses.

Purpose of Study

To visualize immune cells in a physiologically relevant 3D context.
To track the migration patterns of these cells.
To enhance understanding of immune cell interactions.

Methods Used

Preparation of a collagen matrix for cell embedding.
Application of light-sheet microscopy for imaging.
Tracking of cell movement within the 3D matrix.
Utilization of chilled solutions to maintain cell viability.

Main Results

Successful visualization of immune cells in 3D collagen matrices.
Demonstrated efficient tracking of cell migration.
Highlighted the advantages of light-sheet microscopy.
Provided a reproducible protocol for researchers.

Conclusions

This protocol offers a valuable tool for studying immune cell dynamics.
Light-sheet microscopy significantly enhances imaging capabilities.
The method can be adapted for various suspension cells.

Frequently Asked Questions

What is light-sheet microscopy?

Light-sheet microscopy is an imaging technique that illuminates a sample with a thin sheet of light, allowing for fast imaging with reduced phototoxicity.

How does this protocol benefit immunology research?

It allows researchers to visualize and track immune cell behavior in a 3D environment, providing insights into their interactions and migration patterns.

Can this method be used for other cell types?

Yes, the protocol can be adapted for various types of suspension cells within a 3D matrix.

What are the advantages of using a collagen matrix?

Collagen matrices provide a physiologically relevant environment that mimics natural tissue, facilitating more accurate studies of cell behavior.

Who demonstrated this procedure?

Dr. Renping Zhao, a post-doc from the lab, demonstrated the procedure.

What is the significance of tracking cell migration?

Tracking cell migration helps researchers understand how immune cells locate and interact with target cells in a 3D space.

Aqui, apresentamos um protocolo para visualizar as células imunes, incorporadas em uma matriz de colágeno (3D) tridimensionais usando microscopia de luz-folha. Este protocolo também elabora como controlar migração celular em 3D. Este protocolo pode ser empregado para outros tipos de suspensão de células na matriz 3D.

Este método pode ajudar a responder a questões-chave no campo da imunologia, como exatamente as células imunes se comportam em um contexto fisiologicamente relevante, como pesquisar células-alvo de forma eficiente em um ambiente tridimensional. A principal vantagem deste método é gerar uma folha de luz muito fina para iluminar apenas o plano focal sem afetar as células fora do plano. Isso permite uma velocidade de aquisição muito rápida com uma redução pronunciada do branqueamento e da fotocitotoxicidade.

Demonstrando este procedimento estará o Dr. Renping Zhao, um pós-doutorado do meu laboratório. Para começar, sob uma capa de cultura de células, transfira 400 microlitros de solução de estoque de colágeno resfriado para um tubo estéril de 1,5 mililitro. Adicione lentamente 50 microlitros de PBS 10X resfriado.

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