Name: quantitative immunohistochemistry of the cellular microenvironment in patient glioblastoma resections
Uploaded: 2017-07-31T15:00:00
Description: Watch this Scientific Journal Video about Quantitative Immunohistochemistry of the Cellular Microenvironment in Patient Glioblastoma Resections at JoVE.com

作者感谢Drs。 Fahad Bafakih和Jim Mandell收购和鉴定患者样本，Garrett F. Beeghly为免疫组化提供帮助，生物制剂和组织研究设施，心血管研究中心组织学核心和弗吉尼亚大学生物分子分析设备协助样本采集，免疫组织化学和成像。

该方案鉴定了福尔马林固定石蜡包埋（FFPE）样品中的细胞组分，如同样的临床患者样本。石蜡包埋允许细胞和组织形态的最佳维持以及具有更好的切片寿命。用于该分析的样品通过弗吉尼亚大学生物原理和组织研究设施进行访问。根据2010年至2013年在弗吉尼亚大学完成肿瘤切除术的胶质母细胞瘤（星形细胞瘤，WHO IV级）的确定性诊断，由神经病理学家选择患者样本，并在该分析之前被去除<sup class...

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我们在这里提出的方法是一种定量的方法来分析使用传统的显色免疫组织化学染色的组织学样品。此类分析的现行方法包括相似的染色方案，随后由独立病理学家进行分级。该方法是可靠的，但是对于许多应用，需要更精确地了解细胞构成，例如更好地理解与肿瘤相关的异质性和用于体外研究的肿瘤的准确重现。 该方案证明了用于定量分析脑肿瘤微环境内胶质细胞群体的直?...

成胶质细胞瘤（GBM），最常见的恶性脑肿瘤，从初级肿瘤主体，其特征在于高度扩散侵入周围健康脑实质1,2。此漫入侵使得肿瘤尤其难以完全切除，并且保持治疗后入侵癌细胞是不可避免的复发2，3，4中的最常见的原因。以前，我们发现抑制弥漫性胶质瘤细胞侵袭是治疗有益的5 ，但是对于有助于GBM入侵的复杂机制知之甚少。肿瘤微环境中，或组织中癌症周围，已牵涉于肿瘤的进展在多种癌症6,7。胶质母细胞瘤肿瘤微环境，关节炎，相对低于特征，是独特复杂的，由多种胶质细胞组成，如星形胶质细胞，小神经胶质细胞和少突胶质细胞，以及细胞外基质，可溶性因子和生物物理因素。在实验上，星形胶质细胞和小胶质细胞已被证明能增加神经胶质瘤进展和侵袭8，9，10，但所有的神经胶质细胞中的天然人脑微环境中的组合物是未知的。 我们以前显示微环境成分可以通过定量分析胶质母细胞瘤微环境的细胞成分并将我们的分析纳入比例风险模型来预测患者存活11 。在这里，我们描述了定量分析方法，用于鉴定胶质母细胞瘤患者肿瘤切除肿瘤体积和相邻区域内胶质细胞的数量秒。我们使用显色免疫组织化学来鉴定胶质细胞群和ImageJ，以分析每个胶质细胞群体的染色百分比。评估百分比覆盖率创建一个简单的测量来确定细胞的形态学差异，特别是那些受癌细胞相互作用影响的细胞。以前的定量组织病理学染色的研究使用标准染色，如苏木精和曙红12或Masson的三色染色体13 ，其不利用基于抗体的免疫组织化学染色的特异性。我们的方法被开发直接量化胶质母细胞瘤患者肿瘤切除术中的胶质细胞群，我们旨在阐明复杂胶质母细胞瘤微环境。

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			<td height="42" style="height:42px;width:216px;">Xylene</td>
			<td style="width:71px;">Fisher Chemical</td>
			<td style="width:119px;">X3P</td>
			<td style="width:167px;"></td>
		</tr>
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			<td height="21" style="height:21px;width:216px;">Ethanol</td>
			<td style="width:71px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">High pH antigen unmasking solution</td>
			<td style="width:71px;">Vector Labs</td>
			<td>H-3301</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">TBS</td>
			<td style="width:71px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Triton-X</td>
			<td style="width:71px;">Amresco</td>
			<td style="width:119px;">9002-93-1</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Horse serum</td>
			<td style="width:71px;"></td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Anti-ALDH1L1&#160;</td>
			<td style="width:71px;">abcam&#160;</td>
			<td style="width:119px;">ab56777</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Anti-Iba1&#160;</td>
			<td style="width:71px;">abcam&#160;</td>
			<td style="width:119px;">ab5076</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Anti-Oligodendrocyte Specific Protein1&#160;</td>
			<td style="width:71px;">abcam&#160;</td>
			<td style="width:119px;">ab53041</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">ImmPRESS anti-goat</td>
			<td style="width:71px;">Vector Labs</td>
			<td style="width:119px;">MP-7405</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">ImmPRESS Universal (anti-mouse/rabbit)</td>
			<td style="width:71px;">Vector Labs</td>
			<td style="width:119px;">MP-7500</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Hydrogen peroxide</td>
			<td style="width:71px;">Sigma Aldrich</td>
			<td style="width:119px;">216763</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">ImmPACT DAB substrate</td>
			<td style="width:71px;">Vector Labs</td>
			<td style="width:119px;">SK-4105</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Hematoxylin counterstain</td>
			<td style="width:71px;">ThermoScientific</td>
			<td style="width:119px;">72404</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Histochoice Mounting Media</td>
			<td style="width:71px;">Amresco</td>
			<td style="width:119px;">H157-475</td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Aperio Scanscope</td>
			<td style="width:71px;">Leica Biosystems</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="63">
			<td height="63" style="height:63px;width:216px;">Image Scanscope</td>
			<td style="width:71px;">Leica Biosystems</td>
			<td style="width:119px;"></td>
			<td></td>
		</tr>
		<tr height="84">
			<td height="84" style="height:84px;width:216px;">Super HT PAP Pen</td>
			<td style="width:71px;">Research Products International</td>
			<td style="width:119px;">195506</td>
			<td></td>
		</tr>
	</tbody>
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quantitative immunohistochemistry of the cellular microenvironment in patient glioblastoma resections

随着对肿瘤微环境的兴趣越来越大，我们着手开发一种方法，专门确定患者胶质母细胞瘤，最致命和最具侵袭性脑癌患者样本中的微环境成分。不仅定量方法有利于准确描述患病组织，而且还可能有助于更精确的预后，诊断和组织工程系统和替代品的发展。在胶质母细胞瘤中，胶质细胞，如小神经胶质细胞和星形胶质细胞，与病理学家分级的不良预后独立相关。然而，这些细胞和其他神经胶质细胞成分的状态在定量上尚未得到很好的描述。由于标记这些胶质细胞的大过程，这可能是困难的。此外，大多数组织学分析集中在整体组织样本或仅在大部分肿瘤内，而不是基于区域的划分量化在高度异质的组织。在这里，我们描述了一种用于鉴定和定量分析来自成胶质细胞瘤患者的肿瘤切除肿瘤体积和相邻区域内的神经胶质细胞群体的方法。我们使用显色免疫组织化学来鉴定患者肿瘤切除术和ImageJ中的胶质细胞群，以分析每个胶质细胞群体的染色百分比。通过这些技术，我们能够更好地描述胶质瘤肿瘤微环境区域的神经胶质细胞。

对于这一分析，我们的肿瘤切除术中的两个兴趣区域 - 主要肿瘤体积和相邻区域，主要由具有扩散性侵袭性癌细胞的健康组织组成（ 图1A ， 1B ） - 通过合成神经病理学家在苏木精和伊红染色患者样本。在每个患者样品中，通过显色免疫组织化学鉴定了星形胶质细胞（ 图1C ），小胶质细胞（ 图1D</str...

Watch this Scientific Journal Video about Quantitative Immunohistochemistry of the Cellular Microenvironment in Patient Glioblastoma Resections at JoVE.com

Quantitative Immunohistochemistry of the Cellular Microenvironment in Patient Glioblastoma Resections

患者胶质母细胞瘤切除中细胞微环境的定量免疫组织化学

该方案被开发用于使用显色免疫组织化学和ImageJ定量鉴定成胶质细胞瘤患者切除中的肿瘤微环境成分。

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特发性脊柱侧弯的使用蜂窝介电谱的预后预测细胞为基础的检测协议

This protocol details the experimental and analytical procedure for a cell-based assay developed in our laboratory as a functional test to predict the ...

Molecular Profiling of the Invasive Tumor Microenvironment in a 3-Dimensional Model of Colorectal Cancer Cells and Ex vivo Fibroblasts

Molecular Profiling of the Invasive Tumor Microenvironment in a 3-Dimensional Model of Colorectal Cancer Cells and Ex vivo Fibroblasts

在结直肠癌细胞的三维模型的入侵肿瘤微环境的分子剖析和<em&gt;体外</em&gt;成纤维细胞

Invading colorectal cancer (CRC) cells have acquired the capacity to break free from their sister cells, infiltrate the stroma, and remodel the ...

Tumor Treating Field Therapy in Combination with Bevacizumab for the Treatment of Recurrent Glioblastoma

肿瘤治疗领域的治疗与联合贝伐单抗治疗复发性胶质母细胞瘤的治疗

A novel device that employs TTF therapy has recently been developed and is currently in use for the treatment of recurrent glioblastoma (rGBM). It was FDA ...

A "Patient-Like" Orthotopic Syngeneic Mouse Model of Hepatocellular Carcinoma Metastasis

“以病人为像”肝癌原位同源小鼠模型的癌转移

The majority of cancer-related deaths are caused by the metastasis of the cancer rather than the primary tumor itself. Yet, the underlying mechanisms of ...

A Human Glioblastoma Organotypic Slice Culture Model for Study of Tumor Cell Migration and Patient-specific Effects of Anti-Invasive Drugs

人类成胶质细胞瘤器官型切片培养模型，用于研究肿瘤细胞迁移和抗创伤药物的患者特异性作用

Glioblastoma (GBM) continues to carry an extremely poor clinical prognosis despite surgical, chemotherapeutic, and radiation therapy. Progressive tumor ...

Generation of Microtumors Using 3D Human Biogel Culture System and Patient-derived Glioblastoma Cells for Kinomic Profiling and Drug Response Testing

Microtumors使用3D人体Biogel培养体系和病人源性胶质母细胞瘤的Kinomic剖析和药物反应测试的生成

The use of patient-derived xenografts for modeling cancers has provided important insight into cancer biology and drug responsiveness. However, they are ...

Extended Time-lapse Intravital Imaging of Real-time Multicellular Dynamics in the Tumor Microenvironment

实时多细胞动力学延长时间推移活体成像在肿瘤微环境

In the tumor microenvironment, host stromal cells interact with tumor cells to promote tumor progression, angiogenesis, tumor cell dissemination and ...