Name: murine pancreatic islet isolation
Uploaded: 2007-08-22T00:00:00
Description: Watch this Scientific Journal Video about Murine Pancreatic Islet Isolation at JoVE.com

<p class="jove_step">Collagenase Preparation</p>
<ol>
<li>Weigh out the collagenase into a 50 ml conical and add <strong>isolation buffer</strong> to make the final concentration, as indicated in the table below. Dissolve the collagenase by vortexing.<br /><br /><br />(12 mice + 2 extra mice = 14 mice x 5 ml/mouse = 70 ml total collagenase solution)<br /><br /><br />(70 ml x 0.5 mg/ml or 0.3 mg/ml = 35 mg or 21 mg of collagenase P total)<br /><br /><br />
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		<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>Dithizone</td>
<td>reagent</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Dissolve 5mg Dithizone in 5 ml of DMSO in a 50 ml conical tube. Add 45 ml of normal saline and mix gently. Filter with a 60 cc syringe and a 0.22 "m syringe filter into a new 50 ml tube. Store at 4 OC, until use.  Add 1 ml to each 35 mm counting Grid-plate, when needed.</td>
</tr>
<tr>
<td>PBS</td>
<td>Buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>HBSS</td>
<td>Buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Isolation soln.</td>
<td>Buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>HBSS containing 10 mM of HEPES, 1 mM of MgCl2, 5 mM of Glucose, pH 7.4</td>
</tr>
<tr>
<td>Wash soln.</td>
<td>Buffer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Isolation buffer containing 1mM CaCl2</td>
</tr>
<tr>
<td>Collagenase P</td>
<td>&nbsp;</td>
<td>Boehringer Mannheim-Roche</td>
<td>1213873</td>
<td>&nbsp;</td>
<tr>
<td>Dissection tools</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>Hemostat clamp (n=2), Forceps (n=2), Scissors (for skin incision an for taking out pancreas from adjacent tissue)</td>
</tr>
<tr>
<td>Syringe</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>sterile, 5ml, 1/mouse.</td>
</tr>
<tr>
<td>30 G needles</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>sterile, 1/syringe</td>
</tr>
<tr>
<td>siliconized glass vials w/Teflon cap</td>
<td>&nbsp;</td>
<td>Fisher</td>
<td>213-018-54</td>
<td>sterile, 1/mouse</td>
</tr>
<tr>
<td>large sink strainer</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>sterile</td>
</tr>
<tr>
<td>1000 ml Beaker</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>sterile</td>
</tr>
<tr>
<td>Gauze pads</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>4x4, 2/mouse</td>
</tr>
<tr>
<td>plastic eye dropper</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>sterile</td>
</tr>
<tr>
<td>Mice</td>
<td>Animal</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>
			<div>
			&nbsp;		</div>
		

murine pancreatic islet isolation

Watch this Scientific Journal Video about Murine Pancreatic Islet Isolation at JoVE.com

Murine Pancreatic Islet Isolation

Research

JoVE Journal

Biology

Human Pancreatic Islet Isolation: Part I: Digestion and Collection of Pancreatic Tissue

Management of Type 1 diabetes is burdensome, both to the individual and society, costing over 100 billion dollars annually. Despite the widespread use of ...

Human Pancreatic Islet Isolation: Part II: Purification and Culture of Human Islets

Isolation of Stem Cells from Human Pancreatic Cancer Xenografts

Cancer stem cells (CSCs) have been identified in a growing number of malignancies and are functionally defined by their ability to undergo self-renewal ...

Computer-assisted Large-scale Visualization and Quantification of Pancreatic Islet Mass, Size Distribution and Architecture

The pancreatic islet is a unique micro-organ composed of several hormone secreting endocrine cells such as beta-cells (insulin), alpha-cells (glucagon), ...

Mouse Islet of Langerhans Isolation using a Combination of Purified Collagenase and Neutral Protease

The interrogation of beta cell gene expression and function in vitro has squarely shifted over the years from the study of rodent tumorigenic cell lines ...

A Method for Mouse Pancreatic Islet Isolation and Intracellular cAMP Determination

Uncontrolled glycemia is a hallmark of diabetes mellitus and promotes morbidities like neuropathy, nephropathy, and retinopathy. With the increasing ...

Isolation and Culture of Mouse Primary Pancreatic Acinar Cells

This protocol permits rapid isolation (in less than 1 hr) of murine pancreatic acini, making it possible to maintain them in culture for more than one ...

Isolation of Murine Valve Endothelial Cells

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

Isolation of Murine Coronary Vascular Smooth Muscle Cells

While the isolation and culture of vascular smooth muscle cells (VSMCs) from large vessels is well established, we sought to isolate and culture VSMCs ...

A Simple High Efficiency Protocol for Pancreatic Islet Isolation from Mice

Pancreatic islets, also called the Islets of Langerhans, are a cluster of endocrine cells which produces hormones for glucose regulation and other ...