Name: isolation of mononuclear cells from the central nervous system of rats with eae
Uploaded: 2007-12-04T04:23:00
Description: Watch this Scientific Journal Video about Isolation of Mononuclear Cells from the Central Nervous System of Rats with EAE at JoVE.com

<ol>
<li><span style="font-size: x-small; font-family: arial,helvetica,sans-serif;"> </span>Deeply anesthetize rats. Spray with 70% ethanol and do a cardiac perfusion with PBS for 10 min to remove cells from blood vessels (cut the right atria and perfuse through the left ventricle). </li>
<li>Remove the brain and spinal cord and place in a 50 ml tube containing ice-cold PBS. Cut the brain and spinal cord in a 70 mm cell strainer placed in a 10 cm petri dish containing 10 ml of ice-cold PBS. Press each piece of organ through the cell strainer ...

<ol>
	<li>Beeton, C., Chandy, K. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Induction+and+clinical+scoring+of+chronic-relapsing+experimental+autoimmune+encephalomyelitis.">Induction and clinical scoring of chronic-relapsing experimental autoimmune encephalomyelitis.</a> <em style='font-style: italic'>Journal of Visualized Experiments</em>. <b>5</b>, .</li></ol>

<p class="jove_content">This procedure, as all procedures involving live animals, must be approved by your institution's animal use and care committee.  We recommend that a veterinarian or a veterinary technician be present when performing the first cardiac perfusions to ensure a sufficient level of anesthesia is given to the animal before and during the procedure, and that the animals do not undergo unnecessary pain or distress.</p>

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>PBS, 1x, sterile</td>
<td>Reagent</td>
<td>Gibco / Invitrogen</td>
<td>14190-250</td>
<td>&nbsp;</td>
<tr>
<td>Cell strainer, 70 um</td>
<td>Tool</td>
<td>Fisher</td>
<td>08-771-2</td>
<td>&nbsp;</td>
<tr>
<td>Percoll</td>
<td>Reagent</td>
<td>Sigma</td>
<td>P1644</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>
	

isolation of mononuclear cells from the central nervous system of rats with eae

<p class="jove_content">Whether studying an autoimmune disease directed to the central nervous system (CNS), such as experimental autoimmune encephalomyelitis (EAE, 1), or the immune response to an infection of the CNS, such as poliomyelitis, Lyme neuroborreliosis, or neurosyphilis, it is often necessary to isolate the CNS-infiltrating immune cells.</p>
<p class="jove_content">In this video-protocol we demonstrate how to isolate mononuclear cells (MNCs) from the CNS of a rat with EAE.  The first step of this procedure requires a cardiac perfusion of the rodent with a saline solution to ensure that no blood remains in the blood vessels irrigating the CNS. Any blood contamination will artificially increase the number of apparent CNS-infiltrating MNCs and may alter the apparent composition of the immune infiltrate. We then demonstrate how to remove the brain and spinal cord of the rat for subsequent dilaceration to prepare a single-cell suspension. This suspension is separated on a two-layer Percoll gradient to isolate the MNCs. After washing, these cells are then ready to undergo any required procedure. </p>
<p class="jove_content">Mononuclear cells isolated using this procedure are viable and can be used for electrophysiology, flow cytometry (FACS), or biochemistry. If the technique is performed under sterile conditions (using sterile instruments in a tissue culture hood) the cells can also be grown in tissue culture medium. A given cell population can be further purified using either magnetic separation procedures or a FACS.</p>

Watch this Scientific Journal Video about Isolation of Mononuclear Cells from the Central Nervous System of Rats with EAE at JoVE.com

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

<p class="jove_content">In this video we demonstrate how to isolate mononuclear cells from the central nervous system of rats with experimental autoimmune encephalomyelitis. </p>

Whether studying an autoimmune disease directed to the central nervous system (CNS), such as experimental autoimmune encephalomyelitis (EAE, 1), or the ...

Research

JoVE Journal

Biology

Isolation and Analysis of Hematopoietic Stem Cells from the Placenta

Hematopoietic stem cells (HSCs) have the ability to self-renew and generate all cell types of the blood lineages throughout the lifetime of an individual. ...

<em>Homarus Americanus</em> Stomatogastric Nervous System Dissection

Homarus Americanus Stomatogastric Nervous System Dissection

With the goal of understanding how nervous systems produce activity and respond to the environment, neuroscientists turn to model systems that exhibit the ...

<em>Cancer Borealis</em> Stomatogastric Nervous System Dissection

Cancer Borealis Stomatogastric Nervous System Dissection

The stomatogastric ganglion (STG) is an excellent model for studying cellular and network interactions because it contains a relatively small number of ...

Isolation of Retinal Stem Cells from the Mouse Eye

The adult mouse retinal stem cell (RSC) is a rare quiescent cell found within the ciliary epithelium (CE) of the mammalian eye1,2,3. The CE is made up of ...

Isolation and Culture of Cells from the Nephrogenic Zone of the Embryonic Mouse Kidney

Embryonic development of the kidney has been extensively studied both as a model for epithelial-mesenchymal interaction in organogenesis and  to gain ...

Isolation of Murine Valve Endothelial Cells

Isolering af murint Ventil endotelceller

Normal valve structures consist of stratified layers of specialized extracellular matrix (ECM) interspersed with valve interstitial cells (VICs) and ...

Isolation and Primary Culture of Mouse Aortic Endothelial Cells

Isolering og primær kultur af muse aortaendotelceller

The vascular endothelium is essential to normal vascular homeostasis. Its dysfunction participates in various cardiovascular disorders. The mouse is an ...

Isolation of Myoepithelial Cells from Adult Murine Lacrimal and Submandibular Glands

Isolering af Myoepitelial celler fra voksne murine lacrimal og submandibulære kirtler

The lacrimal gland (LG) is an exocrine tubuloacinar gland that secretes an aqueous layer of tear film. The LG epithelial tree is comprised of acinar, ...

Isolation and Culture of Primary Aortic Endothelial Cells from Miniature Pigs

Isolation og kultur af primære aorta endotelceller fra miniature grise

Xenotransplantation is a promising way to resolve the shortage of human organs for patients with end-stage organ failure, and the pig is considered as a ...

Isolation of High Quality Murine Atrial and Ventricular Myocytes for Simultaneous Measurements of Ca<sup>2+</sup> Transients and L-Type Calcium Current

Isolation of High Quality Murine Atrial and Ventricular Myocytes for Simultaneous Measurements of Ca2+ Transients and L-Type Calcium Current

Isolering af murin atriale og ventrikulære myocytter af høj kvalitet til samtidige målinger af<sup>Ca2+</sup> transienter og L-type calciumstrøm

Mouse models play a crucial role in arrhythmia research and allow studying key mechanisms of arrhythmogenesis including altered ion channel function and ...