Name: Induction and Clinical Scoring of Chronic-Relapsing Experimental Autoimmune Encephalomyelitis (Video) | JoVE Journal
Uploaded: 2007-07-04T00:00:00.000+00:00
Duration: 26 min 48 s
Description: Learn to induce and clinically score chronic-relapsing experimental autoimmune encephalomyelitis in DA rats. This method provides a reliable multiple sclerosis model with standardized evaluation for neuroscience research.

ai_agent_IP

ai_agent

maintenance

instantTrialDurationMinutes

isBiopharma

enableNewMyJoveSectionDesign

educationFeatures

seoMetaApi

ai_copilot_institution_blacklist

ai_chat_institution_blacklist

freeAccessWindow

ssoRoleCapture

scheduledMaintenanceBanner

joveLabsFeatures

disable_autoplay_for_bots

extended_trial_duration

5 minutes

playerTrialDesktop

Induction and monitoring of chronic-relapsing EAE
The emulsion is a 1:1 mixture of antigen in aqueous solution added to supplemented complete Freund's adjuvant. It is very important to add the antigen to the adjuvant and not the other way round!!!
There is a lot of waste when making and injecting an emulsion. Always prepare 1.5 or 2 times more than what you need.
Use autoclaved mortar, pestle, glass syringes, and bridges. All plastics should be sterile. The preparation can be done on a regular laboratory bench.
1.&nbsp;Preparation of the supplemented adjuvant
Weigh 30 mg of Mycobacterium tuberculosis (Difco catalog # 231141) and place into a mortar. Make into thin powder without pressing too hard to avoid breaking the bacteria. Add 10 ml of complete Freund's adjuvant H37Ra (Difco catalog # 231131) and mix. This supplemented adjuvant now contains 4 mg/ml Mycobacterium tuberculosis. Transfer to a tube.
2. Preparation of the spinal cord homogenate
Collect spinal cords from DA rats (age/gender indifferent) and store frozen at -80&deg;C. Weigh the frozen spinal cords to have enough to mix 1:1 (weight:volume) with the supplemented adjuvant. Mince the spinal cords as fine as possible with a razor blade, transfer to the mortar used in section 1, and make a paste with the pestle.
3.&nbsp;Preparation of the emulsion
<ol>
<li>Place supplemented complete Freund's adjuvant in a tube. Vortex at high speed. Add the spinal cord homogenate drop by drop while vortexing. When all of the spinal cord homogenate is added, vortex for 5 more minutes. The mixture should turn light pink. </li>
<li>Put the emulsion in a 5 ml glass syringe and link it to another 5 ml glass syringe using a 18G bridge (Fisher catalog # 14-825-17L). Send the emulsion from a syringe to the other until it becomes hard (5-10 min). If more than 5 ml of emulsion is prepared, use several 5 ml syringes DO NOT use larger syringes. </li>
<li>The emulsion can be stored in the syringes at 4&deg;C for 3 weeks. I recommend preparing it at least 12 hours in advance to check that it does not break down. It should remain thick and not separate in 2 phases. The color will change overnight to a light yellow or beige.</li>
</ol>
4.&nbsp;Immunization of the rats
<ol>
<li>Mix the emulsion in the syringes for a few minutes and transfer to the syringe used for the injection. Inject 200 &micro;l subcutaneously at the very base of the tail using 23G needles and 3 ml Luer-Lock syringes under short-term anesthesia. </li>
<li>The recipients are 7-9 weeks old female DA rats. We get our rats from Harlan-Sprague Dawley. </li>
<li>Rats should be observed twice daily and weighed daily. Clinical signs are expected 7-15 days after injection of the emulsion. </li>
</ol>
Clinical scoring:
0: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; no disease
0.5: &nbsp;&nbsp;&nbsp;&nbsp; distal limp tail
1: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; limp tail
2: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; mild paraparesis, ataxia
3: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; moderate paraparesis, the rats trips from time to time
3.5: &nbsp;&nbsp;&nbsp;&nbsp; one hind limb is paralyzed, the other moves
4: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; complete hind limb paralysis
5: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; complete hind limb paralysis and incontinence
6: &nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp; moribund, difficulty breathing, does not eat or drink. Euthanize immediately.

CB and KGC are co-founders and consultants for Airmid Inc.

There are a few important considerations for this protocol.
Genetics of DA rats will differ slightly depending on which breeder they are bought from. These differences may increase or decrease the susceptibility of rats to EAE induction. If your rats are more susceptible to EAE you will want to reduce the strength of the immunization by using non-supplemented complete Freund's adjuvant or even incomplete Freund's adjuvant. You may also consider reducing the amount of emulsion injected or dilu...

<table><tbody><tr><td>Complete Freund&amp;rsquo;s Adjuvant</td><td>Reagent</td><td>Fisher Scientific</td><td>DF 311-360-5</td><td>Manufactured by Difco</td></tr><tr><td>Mycobacterium tuberculosis H37Ra</td><td>Reagent</td><td>Fisher Scientific</td><td>DF3114-33-8</td><td>Manufactured by Difco</td></tr><tr><td>DA rat</td><td>Animal</td><td>Harlan Laboratories</td><td></td><td>Females, 7-9 weeks old.</td></tr><tr><td>Glass syringes - 5 ml</td><td>Tool</td><td>Fisher Scientific</td><td>14-823-10 B</td><td></td></tr><tr><td>Metal bridge - 18G</td><td>Tool</td><td>Fisher Scientific</td><td>14-825-17L</td><td></td></tr><tr><td>Luer Lok plastic syringes - 3 ml</td><td>Tool</td><td>BD Biosciences</td><td>309585</td><td></td></tr><tr><td>Needles - 20G 1 1/2</td><td>Tool</td><td>Fisher Scientific</td><td>14-826-5C</td><td></td></tr></tbody></table>

induction and clinical scoring of chronic-relapsing experimental autoimmune encephalomyelitis

Induction and Clinical Scoring of Chronic-Relapsing Experimental Autoimmune Encephalomyelitis

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by demyelination and glial scaring in areas disseminated in the brain and spinal cord. These lesions alter nerve conduction and induce the disabling neurological deficits that vary with the location of the demyelinated plaques in the CNS (e.g. paraparesis, paralysis, blindness, incontinence).
Experimental autoimmune encephalomyelitis (EAE) is a model for MS. EAE was first induced accidentally in humans during vaccination against rabies, using viruses grown on rabbit spinal cords. Residues of spinal injected with the inactivated virus induced the CNS disease. Following these observations, a first model of EAE was described in non-human primates immunized with a CNS homogenate by Rivers and Schwenther in 1935. EAE has since been generated in a variety of species and can follow different courses depending on the species/strain and immunizing antigen used. For example, immunizing Lewis rats with myelin basic protein in emulsion with adjuvant induces an acute model of EAE, while the same antigen induces a chronic disease in guinea pigs.
The EAE model described here is induced by immunizing DA rats against DA rat spinal cord in emulsion in complete Freund's adjuvant. Rats develop an ascending flaccid paralysis within 7-14 days post-immunization. Clinical signs follow a relapsing-remitting course over several weeks. Pathology shows large immune infiltrates in the CNS and demyelination plaques. Special considerations for taking care for animals with EAE are described at the end of the video.

This video demonstrates the induction and clinical scoring of an animal model of multiple sclerosis: chronic-relapsing experimental autoimmune encephalomyelitis in DA rats. The disease, induced by immunizing rats with an emulsion containing whole rat spinal cord and complete Freund's adjuvant, presents clinical signs resembling the human disease.

Multiple sclerosis (MS) is a chronic inflammatory disease of the central nervous system (CNS) that commonly affects young adults. It is characterized by ...

I am Christine Beaton. I'm an assistant researcher in George Chen's lab in the Department of Physiology and Biophysics of the University of California at Irvine. And what I'm going to show you in the next few days is how to induce and monitor chronic relapsing EAE, which stands for Experimental Autoimmune Encephalomyelitis in DA rats.This is an animal model for multiple sclerosis, or should I say, one of the animal models for multiple sclerosis. And it is characterized by an ascending flacid paralysis in those animals and histological characteristics that resemble the human disease. So this is going to take several days.So the first day I'm going to show you how to prepare the genic emulsion with naive red spinal cord homogenate in complete trans adjuvant. The second day, I'm going to show you how to immunize the rats to UCAE, and then we're going to wait between seven and 15 days to see the first clinical signs in those animals, at which point I'm going to show you the different stages of progression of EAE in different animal. So I'm first going to show you all the reagents and material that are needed to prepare the emulsion.So what I'm using is complete front adjuvant H 37 ra,

induction-clinical-scoring-chronic-relapsing-experimental-autoimmune

Research

JoVE Journal

Biology

In this video we demonstrate how to isolate mononuclear cells from the central nervous system of rats with experimental autoimmune encephalomyelitis.

Isolation of Mononuclear Cells from the Central Nervous System of Rats with EAE

The video demonstrates the induction of experimental autoimmune encephalomyelitis in a mouse model by injecting a combination of myelin oligodendrocyte glycoproteins and inactivated Mycobacterium tuberculosis. The model enables researchers to study disease mechanisms and test therapeutic interventions.

Induction of Experimental Autoimmune Encephalomyelitis in a Mouse Model

In this video, a mouse model of multiple sclerosis is established using experimental autoimmune encephalomyelitis (EAE). The mouse is injected with an emulsion containing a neuronal peptide and an adjuvant, triggering an immune response that activates autoreactive T cells. This is followed by an injection of pertussis toxin (PTX) to increase blood-brain barrier permeability, allowing the autoreactive T-cells to infiltrate the brain. This infiltration initiates a signaling cascade that...

This manuscript describes the methods for induction and scoring of the experimental autoimmune encephalomyelitis (EAE) model, together with the assessment of immune cell distribution and mRNA cytokine levels in lymph nodes, spleen, blood and spinal cord using flow cytometry and quantitative PCR, respectively, at various disease phases.

Induction of Experimental Autoimmune Encephalomyelitis in Mice and Evaluation of the Disease-dependent Distribution of Immune Cells in Various Tissues

Here, we present protocols to investigate impairment of the neurovascular unit during experimental autoimmune encephalomyelitis in vivo. We specifically address how to determine blood-brain barrier permeability and gelatinase activity involved in leukocyte migration across the glia...

Visualizing Impairment of the Endothelial and Glial Barriers of the Neurovascular Unit during Experimental Autoimmune Encephalomyelitis In Vivo

This manuscript presents a protocol to induce active experimental autoimmune encephalomyelitis (EAE) in mice. A method for the isolation and characterization of the infiltrated lymphocytes in the central nervous system (CNS) is also presented to show how lymphocytes are involved in the development of CNS autoimmune disease.

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

The present protocol describes the induction of experimental autoimmune encephalomyelitis in a mouse model using myelin oligodendrocyte glycoprotein and monitoring the disease process using a clinical scoring system. Experimental autoimmune encephalomyelitis-related symptoms are analyzed using mouse femur micro-computed tomography analysis and open field test to assess the disease process comprehensively.

Induction and Diverse Assessment Indicators of Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) serves as an animal model of multiple sclerosis. This article describes an approach for scoring spinal cord inflammation, demyelination, and axonal injury in EAE. Additionally, a method to quantify soluble neurofilament light levels in the mice serum is presented, facilitating the assessment of axonal injury in live mice.

Scoring Central Nervous System Inflammation, Demyelination, and Axon Injury in Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis is one of the most widely used murine models of multiple sclerosis. In the current protocol, C57BL/6J mice of both sexes are immunized with myelin oligodendrocyte glycoprotein peptide, resulting mainly in ascending paresis of the tail and limbs. Here we discuss the protocol of EAE induction and evaluation.

Modeling Multiple Sclerosis in the Two Sexes: MOG35-55-Induced Experimental Autoimmune Encephalomyelitis

Experimental autoimmune encephalomyelitis (EAE) is an animal model of multiple sclerosis (MS), which shares with the human disease a robust humoral autoimmune response. Here, we report a simple and flexible ELISA protocol to quantify autoantibodies in the serum of EAE immunized mice.

Quantification of Autoreactive Antibodies in Mice upon Experimental Autoimmune Encephalomyelitis

Learn to induce and clinically score chronic-relapsing experimental autoimmune encephalomyelitis in DA rats. This method provides a reliable multiple sclerosis model with standardized evaluation for neuroscience research.

Induction and Clinical Scoring of Chronic-Relapsing Experimental Autoimmune Encephalomyelitis

Explore More Videos