Intravital Imaging of Intraepithelial Lymphocytes in Murine Small Intestine

Luo Jia; Karen  L. Edelblum

doi:10.3791/59853

Imágenes intravitales de linfocitos intraepiteliales en el intestino pequeño murino

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Immunology and Infection

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JoVE Journal Immunology and Infection

Intravital Imaging of Intraepithelial Lymphocytes in Murine Small Intestine

Imágenes intravitales de linfocitos intraepiteliales en el intestino pequeño murino

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08:00 min

June 24, 2019

DOI: 10.3791/59853-v

Luo Jia¹, Karen L. Edelblum¹

¹Center for Immunity and Inflammation, Department of Pathology, Immunology and Laboratory Medicine,Rutgers New Jersey Medical School

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Overview

This article describes a method for visualizing GFP-labeled 纬未 IELs through intravital imaging of the murine small intestine using inverted spinning disk confocal microscopy. This technique allows for the tracking of live cells within the mucosa for up to 4 hours, facilitating the investigation of various intestinal immune-epithelial interactions.

Key Study Components

Area of Science

Neuroscience
Immunology
Cell Biology

Background

Intravital microscopy provides insights into cellular interactions within the intestinal epithelium.
The technique allows for high-speed, high-resolution imaging in real-time.
Live imaging can enhance understanding of mucosal immunity and epithelial biology.
Challenges include mastering surgical techniques and optimal mouse positioning.

Purpose of Study

To visualize live immune cells in the intestinal mucosa.
To investigate immune-epithelial interactions in real-time.
To enhance understanding of cellular dynamics in mucosal immunity.

Methods Used

Intravital imaging using inverted spinning disk confocal microscopy.
Injection of Hoechst 33342 Dye Solution into the retro-orbital venous sinus.
Monitoring of live cells for up to 4 hours.
Use of transgenic mice expressing enhanced green fluorescent protein (EGFP).

Main Results

Successful visualization of 纬未 IELs in the murine small intestine.
Real-time tracking of cellular interactions within the mucosa.
Insights into the dynamics of immune cell interactions.
Demonstration of the technique's potential for studying mucosal immunity.

Conclusions

The method provides a powerful tool for studying live cell interactions in the intestine.
It enhances understanding of the role of immune cells in mucosal biology.
Further applications could extend to cancer biology and other areas.

Frequently Asked Questions

What is intravital microscopy?

Intravital microscopy is a technique that allows for the visualization of live cells within a living organism, providing insights into cellular dynamics.

How long can cells be tracked using this method?

Cells can be tracked for up to 4 hours using this imaging technique.

What is the significance of using GFP-labeled cells?

GFP labeling allows for the visualization of specific cell types, enabling researchers to study their behavior and interactions in real-time.

What challenges are associated with this technique?

Challenges include mastering the surgical technique and ensuring optimal positioning of the mouse for image acquisition.

What insights can be gained from this method?

This method provides insights into mucosal immunity, epithelial biology, and cellular interactions within the intestine.

What type of mice are used in this study?

Transgenic mice expressing enhanced green fluorescent protein (EGFP) are used for visualization.

Describimos un método para visualizar los IEL con la etiqueta GFP utilizando imágenes intravitales del intestino delgado murina mediante microscopía confocal de disco giratorio invertido. Esta técnica permite el seguimiento de células vivas dentro de la mucosa hasta 4 h y se puede utilizar para investigar una variedad de interacciones inmunoepiteliales intestinales.

La microscopía intravital de la pequeña mucosa intestinal puede proporcionar información sobre la dinámica espaciotemporal de las interacciones celulares dentro del epitelio entre diferentes tipos de células inmunitarias, o entre estos dos compartimentos. La principal ventaja de esta técnica es que permite la adquisición de imágenes de alta velocidad y alta resolución dentro de la mucosa intestinal en tiempo real. Las imágenes en vivo del intestino pueden proporcionar información adicional sobre las interacciones celulares, o la señalización celular implicada en la inmunidad a la mucosa, la biología epitelial o la biología del cáncer.

Este es un método desafiante que requiere la práctica de la técnica quirúrgica, así como la paciencia y el posicionamiento óptimo del ratón para la adquisición de imágenes. Después de confirmar la falta de respuesta al pellizco en un ratón transgénico anestesiado, expresando una proteína fluorescente verde mejorada, o receptor de células T de EGFP, Gamma-delta, utilice una jeringa de tuberculina para inyectar lentamente 200 microlitros de solución de tinte Hoechst 33342 recién preparada en el seno venoso retro-orbital. De uno a dos minutos después de la inyección, aplicar un ung en los ojos del animal para evitar que se sequen.

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