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

Rouven Schoppmeyer; Renping Zhao; Markus Hoth; Bin Qu

doi:10.3791/57651

İnsan bağışıklık hücreleri üç boyutlu görselleştirme için ışık sayfalık mikroskobu

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

İnsan bağışıklık hücreleri üç boyutlu görselleştirme için ışık sayfalık mikroskobu

Full Text

8,616 Views

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.

Burada, bağışıklık hücreleri ışık sayfalık mikroskobu kullanarak üç boyutlu (3D) kolajen dizey içinde gömülü görselleştirmek için bir iletişim kuralı mevcut. Bu iletişim kuralı da hücre göç 3D iz sürmeyi ayrıntılandırdığı. Bu iletişim kuralı 3D Matris hücrelerinde süspansiyon diğer türleri için istihdam edilebilir.

Bu yöntem, immünoloji alanında, bağışıklık hücrelerinin fizyolojik olarak ilgili bir bağlamda tam olarak nasıl davrandığı, hedef hücrelerin üç boyutlu bir ortamda verimli bir şekilde nasıl aranacağı gibi temel soruların yanıtlanmasına yardımcı olabilir. Bu yöntemin temel avantajı, düzlem dışı hücreleri etkilemeden sadece odak düzlemini aydınlatmak için çok ince bir ışık tabakası oluşturmaktır. Bu, ağartma ve fotositotoksisitenin belirgin bir şekilde azaltılması ile çok hızlı bir alım hızı sağlar.

Bu prosedürü gösteren, laboratuvarımdan bir doktora sonrası olan Dr.Renping Zhao olacak. Başlamak için, bir hücre kültürü başlığı altında, 400 mikrolitre soğutulmuş kollajen stok çözeltisini steril 1.5 mililitrelik bir tüpe aktarın. Yavaşça 50 mikrolitre soğutulmuş 10X PBS ekleyin.

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