Flow Cytometric Analysis of Lymphocyte Infiltration in Central Nervous System during Experimental Autoimmune Encephalomyelitis

Zhe Ji; Chenghua Zhou; Hongshen Niu; Jinshen Wang; Lei Shen

doi:10.3791/61050

Análise Citométrica de Fluxo de Infiltração linfócito no Sistema Nervoso Central durante Encefalo...

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Immunology and Infection

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JoVE Journal Immunology and Infection

Flow Cytometric Analysis of Lymphocyte Infiltration in Central Nervous System during Experimental Autoimmune Encephalomyelitis

Análise Citométrica de Fluxo de Infiltração linfócito no Sistema Nervoso Central durante Encefalomielite Autoimune Experimental

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09:01 min

November 17, 2020

DOI: 10.3791/61050-v

Zhe Ji^1,2, Chenghua Zhou¹, Hongshen Niu³, Jinshen Wang¹, Lei Shen³

¹Translational Medicine Research Center, Ruijin Hospital North,Shanghai Jiao Tong University School of Medicine, ²Department of Microbiology and Immunology,Tongji University School of Medicine, ³Shanghai Institute of Immunology, Shanghai Key Laboratory for Tumor Microenvironment and Inflammation,Shanghai Jiao Tong University School of Medicine

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Overview

This manuscript presents a protocol to induce active experimental autoimmune encephalomyelitis (EAE) in mice. The method allows for the isolation and characterization of infiltrated lymphocytes in the central nervous system (CNS), highlighting their role in CNS autoimmune disease.

Key Study Components

Area of Science

Neuroscience
Immunology
Autoimmune diseases

Background

Active experimental autoimmune encephalomyelitis (EAE) is a model for studying CNS autoimmune diseases.
Lymphocytes play a crucial role in the development of these diseases.
Understanding lymphocyte behavior can provide insights into immune responses in the CNS.
This protocol can be adapted for other CNS disease models.

Purpose of Study

To develop a reliable method for inducing EAE in mice.
To isolate and analyze lymphocytes infiltrating the CNS.
To enhance understanding of immune cell functions in CNS autoimmune diseases.

Methods Used

Subcutaneous injection of emulsified MOG 35-55 in complete Freund's adjuvant.
Intraperitoneal injection of pertussis toxin to enhance EAE induction.
Isolation of lymphocytes from the CNS for detailed analysis.
Cell-by-cell examination of lymphocyte characteristics and functions.

Main Results

Successful induction of EAE in C57BL/6 mice.
Isolation of infiltrated lymphocytes from the CNS.
Demonstration of lymphocyte involvement in CNS autoimmune disease.
Potential application of the protocol to other CNS models.

Conclusions

The protocol provides a valuable tool for studying CNS autoimmune diseases.
Insights gained can inform therapeutic strategies targeting immune responses.
Further research can expand the applicability of this method to other conditions.

Frequently Asked Questions

What is EAE?

EAE stands for experimental autoimmune encephalomyelitis, a model used to study autoimmune diseases of the CNS.

How are lymphocytes isolated from the CNS?

Lymphocytes are isolated using specific protocols that involve tissue dissociation and cell sorting techniques.

Can this protocol be used for other diseases?

Yes, the protocol can be adapted for studying other CNS diseases that involve immune cell infiltration.

What role do lymphocytes play in CNS diseases?

Lymphocytes are key players in the immune response and can contribute to the pathology of CNS autoimmune diseases.

What is the significance of using C57BL/6 mice?

C57BL/6 mice are a commonly used strain in immunology research due to their well-characterized immune system.

Este manuscrito apresenta um protocolo para induzir encefalomielite autoimune experimental ativa (EAE) em camundongos. Um método de isolamento e caracterização dos linfócitos infiltrados no sistema nervoso central (SNC) também é apresentado para mostrar como os linfócitos estão envolvidos no desenvolvimento da doença autoimune cns.

O protocolo que apresentamos aqui será útil para entendermos como os linfócitos estão envolvidos no desenvolvimento da doença autoimune do sistema nervoso central. Com este método, linfócitos no cérebro podem ser estudados em uma célula por célula. Este protocolo também pode ser aplicado a outros modelos e doenças do sistema nervoso central que precisavam analisar as características e funções das células imunes.

Neste vídeo, o protocolo pode facilmente demonstrar como induzir modelo e isolar células únicas no cérebro. Depois de confirmar a falta de resposta ao reflexo do pedal, use uma seringa de um mililitro para injetar subcutâneamente camundongos C57BL/6 anestesiados com 100 microgramas de MOG 35-55 em 100 microlitres de completojuvante de Freund em dois locais diferentes da parte de trás perto do pescoço. No mesmo dia e no segundo dia pós-imunização, injete intraperitoneally os camundongos com 200 microliters de um nanograma por solução de trabalho de toxina de coqueluche microliter.

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