Name: neutrophil isolation protocol
Uploaded: 2008-07-23T22:05:00
Description: Watch this Scientific Journal Video about Neutrophil Isolation Protocol at JoVE.com

<p>资金从美国国立卫生研究院R01 HL56621。 </p>

<p class="jove_title">嗜中性粒细胞的分离协议</p><ol><li>将所有试剂置于室温。 </li><li>收集的5.0毫升离心管中的中性粒细胞隔离媒体。仔细层5.0毫升的血液分离介质，。执行这一步，缓慢而仔细地，并与介质的表面，以避免混合血液和媒体的枪头。 </li><li> 500 35分钟的离心力离心20-25℃。血液分离出6个不同组别：血浆，单核细胞，隔离媒体，中性粒细胞，更加孤立的媒体，以及红血细胞沉淀（图1）。如果不明确，这些?...

<ol>
	<li>Boyum, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Separation+of+leucocytes+from+blood+and+bone+marrow.">Separation of leucocytes from blood and bone marrow.</a> <em style='font-style: italic'>Scand. J. Clin. Invest</em>. <b>21</b>, 77-83 (1968).</li><li>England, J. M., Rowan, R. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+assignment+of+values+to+fresh+blood+used+for+calibrating+automated+blood+cell+counters.">The assignment of values to fresh blood used for calibrating automated blood cell counters.</a> <em style='font-style: italic'>Clin. Lab. Haemat</em>. <b>10</b>, 203-212 (1988).</li><li>Garcia, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparison+of+two+leukocyte+extraction+methods+for+cytomegalovirus+antigenemia+assay.">Comparison of two leukocyte extraction methods for cytomegalovirus antigenemia assay.</a> <em style='font-style: italic'>J. Clin. Microbiol</em>. <b>34</b>, 182-184 (1996).</li><li>Ferrante, A., Thong, Y. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Optimal+conditions+for+simultaneous+purification+of+mononuclear+and+polymorphonuclear+leucocytes+from+human+peripheral+blood+by+the+Ficoll-Hypaque+method.">Optimal conditions for simultaneous purification of mononuclear and polymorphonuclear leucocytes from human peripheral blood by the Ficoll-Hypaque method.</a> <em style='font-style: italic'>J. Immunol. Methods</em>. <b>36</b>, 109-109 (1980).</li></ol>

<p>密度梯度分离法是用来隔离从全血混合使用metrizoate钠和右旋糖酐500人中性粒细胞。这种方法是基于对Boyum（1968年）的单核白细胞分离法修改费兰特和通（1980）中性粒细胞分离。 </p><p>从捐赠者收集后，全血用EDTA，柠檬酸或肝素抗凝。由于它们是短命的，中性粒细胞应收集2-4小时内使用。由分层全血通过离心，密度梯度介质，中性粒细胞层的分离，剩余红​​细胞裂解的过程。细胞，然后洗净，计算，并重新悬?...

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		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
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<tr>
<td>Lymphocyte Poly(R)</td>
<td>Reagent</td>
<td>Cedarlane Labs</td>
<td>&nbsp;</td>
<td>PolymorphprepTM can be used as an alternative</td>
</tr>
<tr>
<td>Hank's Balanced Salt Solution without Calcium Chloride</td>
<td>Reagent</td>
<td>Invitrogen Gibco</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Human Serum Albumin</td>
<td>Reagent</td>
<td>ZLB Bioplasma</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Red Cell Lysis Buffer</td>
<td>Reagent</td>
<td>Roche Diagnostics</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Centrifuge</td>
<td>Tool</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Vortexer</td>
<td>Tool</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>Pasteur Pipettes and bulb</td>
<td>Tool</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<td>&nbsp;</td>
<tr>
<td>PolymorphprepTM</td>
<td>Reagent</td>
<td>Axis-Shield PoC</td>
<td>&nbsp;</td>
<td>Alternative reagent to Lymphocyte-Poly (R)</td>
</tr>
<tr>
<td>Syringe Filter</td>
<td>Other</td>
<td>Millipore</td>
<td>SLGV033RS</td>
<td>Millex-GV, 0.22 &#181;m, PVDF, 33 mm, gamma-sterilizable</td>
</tr>
<tr>
<td>Hank's Balance Salt Solution with Calcium Chloride</td>
<td>Reagent</td>
<td>Invitrogen Gibco</td>
<td>&nbsp;</td>
<td>&nbsp;</td>		</tbody>
		</table>
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neutrophil isolation protocol

<p class="jove_content">中性粒细胞的多形核粒细胞（PMN）在人类之间的第一个细胞到达炎症免疫反应的部位，最丰富的白细胞。由于其关键作用，在炎症，如中性粒细胞功能的运动，细胞因子的产生，的吞噬功能，和肿瘤细胞的作战广泛的研究。为特征的中性粒细胞的特定功能，其他血细胞分离干净，快速，可靠的方法是可取的，尤其是因为在体外研究中性粒细胞是短命的，应在2-4小时收集。在这里，我们展示了一个标准的密度梯度分离法，从全血使用市售分离介质，是一种混合物，钠metrizoate和右旋糖酐500人中性粒细胞隔离。由分层全血通过离心，密度梯度介质，中性粒细胞层的分离，剩余红​​细胞裂解的过程。细胞，然后洗净，计数，并在缓冲区所需浓度的悬浮。正确执行时，此方法已被证明收益率&gt; 95％嗜中性粒细胞&gt; 95％的可行性的样品。</p>

Watch this Scientific Journal Video about Neutrophil Isolation Protocol at JoVE.com

Neutrophil Isolation Protocol

<p class="jove_content">嗜中性粒细胞之间的第一个细胞到达炎症免疫反应的工地上，和其职能和机制，已被广泛地在体外研究。我们展示一个标准密度梯度分离法，隔离人类使用市售的分离介质从整个血液中性粒细胞。</p>

嗜中性粒细胞的分离协议

Neutrophil polymorphonuclear granulocytes (PMN) are the most abundant leukocytes in humans and among the first cells to arrive on the site of inflammatory ...

Research

JoVE Journal

Biology

Protocol for Mosquito Rearing (A. gambiae)

Protocol for Mosquito Rearing A. gambiae

蚊子养育的议定书“（冈比亚按蚊）

This protocol describes mosquito rearing in the insectary. The insectary rooms are maintained at 28&deg;C and ~80% humidity, with a 12 hr. day/night ...

科研

生物学

A Protocol for the Production of KLRG1 Tetramer

KLRG1四聚体的生产协议

Killer cell lectin-like receptor G1 (KLRG1) is a type II transmembrane glycoprotein inhibitory receptor belonging to the C type lectin-like superfamily. ...

DNA-based Fish Species Identification Protocol

以DNA为基础的鱼类物种鉴定协议

We have developed a fast, simple, and accurate DNA-based screening method to identify the fish species present in fresh and processed seafood samples. ...

Protocol for Isolation of Primary Human Hepatocytes and Corresponding Major Populations of Non-parenchymal Liver Cells

协议主要人肝细胞的分离和相应的非实质肝细胞的主要种群

Beside parenchymal hepatocytes, the liver consists of non-parenchymal cells (NPC) namely Kupffer cells (KC), liver endothelial cells (LEC) and hepatic ...

Clock Scan Protocol for Image Analysis: ImageJ Plugins

用于图像分析的时钟扫描协议：ImageJ插件

The clock scan protocol for image analysis is an efficient tool to quantify the average pixel intensity within, at the border, and outside (background) a ...

A Simple High Efficiency Protocol for Pancreatic Islet Isolation from Mice

胰腺胰岛与小鼠分离的简单高效协议

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

The MUB<sub>40</sub> Peptide for Use in Detecting Neutrophil-Mediated Inflammation Events

The MUB40 Peptide for Use in Detecting Neutrophil-Mediated Inflammation Events

mub<sub>40</sub>肽在中性炎症事件检测中的应用

Here, we provide a protocol involving the use of MUB40, a synthesized peptide with the ability to bind glycosylated lactoferrin stored at high ...

Rapid Lipid Droplet Isolation Protocol Using a Well-established Organelle Isolation Kit

利用成熟的细胞隔离试剂盒快速分离脂滴器

Lipid droplets (LDs) are bioactive organelles found within the cytosol of the most eukaryotic and some prokaryotic cells. LDs are composed of neutral ...

An Improved Time- and Labor- Efficient Protocol for Mouse Primary Hepatocyte Isolation

一种改进的小鼠原代肝细胞分离时间和劳动效率方案

Primary hepatocytes are used extensively in liver in vitro research, especially in glucose metabolism studies. A base technique has been adapted based on ...

Neutrophil Lifespan Extension with CLON-G and an <em>In Vitro</em> Spontaneous Death Assay

Neutrophil Lifespan Extension with CLON-G and an In Vitro Spontaneous Death Assay

通过 CLON-G 和 <em>体外</em> 自发死亡测定延长中性粒细胞寿命

The average lifespan of a neutrophil is less than 24 h, which limits basic research on neutrophils and the application of neutrophil studies. Our previous ...