Name: spot the correct tissue every time in multi-tissue blocks
Uploaded: 2015-05-31T14:00:00.000+00:00
Duration: 6 min 53 s
Description: Watch this Scientific Journal Video about SpOT the Correct Tissue Every Time in Multi-tissue Blocks at JoVE.com

作者要感谢布伦特·哈里斯博士提供稿件的严格审查。这些研究在隆巴迪综合癌症中心组织病理与组织共享资源是支持由NIH / NCI资助P30-CA051008部分进行。内容完全是作者的责任，并不一定代表美国国家癌症研究所和美国国立卫生研究院的官方意见。

1.施工现场的<ol><li>添加50毫克牛血清白蛋白（BSA）的1毫升组织标记染料在15ml锥形管或2 ml离心管，涡旋1分钟，或直至完全溶解。 注意:对于这个协议，使用生化级BSA。而其他等级和纯度都没有经过测试，预计品位和纯度不会对现货的成功显著的影响。 </li><li>热9毫升羟乙基在15ml的锥形管在微波中，在30％的功率，持续10秒的增量琼脂糖处理凝胶直到凝胶完全融化。 </li><li>结合熔融加工的凝胶和染料BSA溶液在单一的15毫升锥形管中，并用移液管充分混合。涡的解决方案。 </li><li>放置锥形管在冰箱中几个小时或直至固体。 </li><li>使用长而细，金属铲或探针从锥形管轻轻松开彩色凝胶塞。 </li><li>使用手术刀或刀片来CUt是凝胶横插分成5毫米厚的切片。取出圆端和处置废物（ 图2A）。 </li><li>将凝胶塞部分平面中组织学盒并保持在70％乙醇中1小时。至少3的变化，在24小时内发生变化的70％的乙醇溶液每2-4小时。 </li><li>手动处理通过乙醇系列（1小时每凝胶部分:70％乙醇，80％乙醇，95％乙醇，100％乙醇（3×的变化），然后清除在清除溶液3次1小时的每个（ 例如，脂族烃（二甲苯替代））在这个协议中使用，二甲苯也可以接受），并渗透有熔融石蜡（3×为每1小时），可以手动或以组织processer。 注:在充分脱水在100％乙醇中，染料可能会泄漏从凝胶塞这可能会影响在自动组织处理器其它组织和液体试剂。因此，人工补液强烈建议，但自动化处理EQU盟友有效。 </li><li>嵌入多个处理凝胶切片用标准石蜡包埋方法（ 图2B）。允许块冷却并硬化，然后回复到室温。 </li><li>使用所需尺寸的真皮冲针从嵌入凝胶切片，直至该块被耗尽（ 图2C）除去斑核。单个块可提供高达20×2平方毫米的核心。存储内核在阴凉处。使用真皮冲头在这个协议中看到图中，在1.5毫米（ 图1D和2D）或2毫米（ 图1B，1C和2C）。 </li></ol> 2.使用斑点东方非阵列块<ol><li>创建组织取向图以指示所希望的位置和所有单独的组织块要被嵌入在一起的身份，和光点的位置。 注:该组织的方向图是一本图文并茂的地图，包括p所有的组织块标有标识符和点的位置的位置物质环境。它可以以电子方式创建，也可以手工绘制。该图应使用任何方法最适合使用的最终产品的技术人员和研究人员来创建。此地图将作为研究者导向，以便样品可以在微观分析容易地识别。在手绘本协议中使用的地图。它描绘了与每片组织和绘制并标记在相同的位置的实际组织块和滑动当场组织块和显微镜载玻片。 </li><li>处理组织片通常，以确保它们保持正确整个票房和处理步骤识别。此协议，处理该组织1小时每中:70％的乙醇，80％乙醇，95％乙醇，100％乙醇（×3的变化），脂族烃（二甲苯代替）（×3的变化），和熔融的组织浸润石蜡在60°C（X3的变化）。 注意:LL组织应按照标准组织学实验室程序进行处理。这可以是可变的，由于各个实验室过程，组织大小和类型，但是这将不会影响到使用点作为定向工具的能力。 </li><li>根据该组织的取向图安排组织片的嵌入站上以正确的分配位置。使用方向组织图作为参考来指导如何安排，当场组织块每块技术员。 注意:在这个协议中嵌入技师把手绘取向图旁边嵌入台和布置在组织片的嵌入站上完全相同的结构所绘制在图上。这确保了所有组织片留在最终产物中正确地识别。这是成功使用多组织块的绝对关键。 </li><li>确保当场核心是身高超过所有组织块被嵌入在该块。 </li><li>根据使用标准嵌入方法组织方向图嵌入的组织块，然后在年底（横向）的地点在合适的位置。根据需要，召开现场会（S）直立钳，直到石蜡凝固，以至于现场（S）也不会摔倒（1-2分钟）。 </li><li>科和染色石蜡切片使用标准方法的地方。 <ol><li>允许部分浮在水浴中于40℃和粘附到带正电荷的载玻片（不带电荷的载玻片尚未经过测试）。干燥的干燥炉的幻灯片，在60℃下进行至少20分钟。 </li><li>将载玻片在自动染色机和加工通过下列试剂:二甲苯（二甲苯1-5分钟，二甲苯2和3-2分钟，每），100％乙醇（2分钟），95％乙醇（1分钟），80 ％乙醇（30秒），70％的乙醇（30秒），自来水（2分钟），苏木（1.5分），自来水（3分钟），酸醇（1分钟），自来水（2分钟），BLuing试剂（1分钟），自来水（2分钟），80％的乙醇（30秒），曙红（1分钟），自来水（10秒），80％乙醇（1分钟），100％乙醇（3变化，每2分钟），二甲苯（3变化，每30秒）。 </li><li>盖上玻璃盖玻片，安装有安装介质的幻灯片。 注:在这个协议中，组织学技术人员在切片5微米的部分上切片的石蜡块。对于这个协议，我们使用一个自动染色机，但平等的结果可以通过手动染色获得。 </li></ol></li></ol> 3.现货在组织芯片（TMA手动工具包） <ol><li>创建一个组织的方向图，如2.1描述和分配点的所需位置（S）。 </li><li>填写TMA模具融化的石蜡在嵌入站。由于模具冷却，嵌入到底当场在TMA模具在由组织定向图所示所需位置的边缘（S）。根据需要，召开现场会（S）直立钳，直到石蜡公顷s凝固足够，现货（S）也不会摔倒（1-2分钟）。 </li><li>根据制造商的指示，并就在数组利润率（ 图2D）点（S）的位置组装TMA内核，并按照标准切片和染色协议。 注:请参见步骤2.6看到在这个协议中使用的切片和染色方法。 </li></ol>

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作者什么都没有透露。

成功制备的斑点和利用需要认真遵守一些技术步骤。羟乙基琼脂糖熔化应缓慢并且在低热完成。在高的热迅速熔化可导致琼脂糖一些击穿不到最佳效果。当切割染料载货插头插入件进行处理，确保每一块的厚度大于4.5毫米，不超过7 mm。的圆形端，因为它们不是均匀5mm的厚度被除去废物。件小于5mm将导致该就有可能被用尽之前由邻近组织材料用尽切片的创建短斑点。为了确保现场出现过块的整体，件需要至少4.5-5毫米。以确保蜡的适当浸润，然而，件应不超过7 mm。当脱水染料片为蜡渗透，染料会稍微浸出件在罗威ř浓度的乙醇。浸出不会影响最终产品，斑点将仍然是强烈的染色和清晰可见。浸出可能会影响其它组织，虽然如此，组织切片不应在同一浴进行处理作为染料片，直到片已达到100％的乙醇。 当把这些点进入摆着块，确保所有其它组织的位置是将现场之前完成。由于现场被渗透与石蜡，收件人块熔化的蜡就可以开始融化当场核是否允许坐太久。先安排所有的组织块，加当场核心，并立即块转移到冷板，以防止斑点的任何熔化。 当场就可以进行修改，以适应大多数组织学实验室的工作流程。染料的颜色可以为每个应用最佳的对比度来改变。红色或黑色​​染料产生的免疫组化染色切片清晰的高对比度，W往往微不足道的红点是不一样的排列肾切片的H＆E染色的幻灯片醒目。对于H＆E幻灯片，绿色或黄色斑点往往会提供更高的对比度。此外，在过程中如热诱导抗原修复免疫组化进程高热，羟乙基琼脂糖融化。当场制备过程中的BSA加入到染料保持在即使经过高温检索方式滑动颜色鲜艳的斑点。如果滑动将不会暴露在高温，波塞可以省略更快点生产。 利用现货的最关键的环节是在创造和维护一个清晰，简明的方位图，并把现场当忠实遵守地图。如果使用得当，现货减少混乱的组织安排，并简化了技术人员和研究人员之间的交流，作为嵌入技术人员可以简单地表示点的位置和Tissu酒店的关系下载到它在地图上。 SPOT是所有步骤的幻灯片准备允许高度一致的部分安装在滑轨，便于下游分析清晰可见。与此相反，以生理标记，光斑在每节清晰可见并且不修改所述组织以任何方式，防止引入假象或梗阻形态的。不同于不对称嵌入组织的方法，当场让技术人员和研究人员组织的安排灵活性的显微镜下，以优化的可视化分析。在的TMA，当场就可以被放置在主阵列外，省去了空格，非对称组件，或信标核心，允许有价值的组织切片的整个阵列组成。当场也可用于指示解剖方向的组织样本（ 例如，远端或近端）。现货对组织样本的结构和过程中很容易和一致取向的简单和低成本的解决方案nalysis多组织块和组织芯片的。

嵌入在一个单一的石蜡块从多个个体的组织样品的能力使得护理和个人之间的容易的侧方比较，消除了幻灯片之间的变异，并降低切片和染色标本的成本和工作量。这些多组织块通常产生含有在非阵列布局从多个个体组织或者组织微阵列（TMA）或石蜡块。样本标识的保养是任何多组织分析的成功是至关重要的。使用的TMA因为它们的发展，提高分析的效率，减少幻灯片之间变化，节省宝贵的组织资源，并减少的时间和实验1成本研究者已经。的TMA的正确取向可以采用多种方法来完成，包括组织核心2-4，芯组的非对称布置3，5（ 例如，CONTRO之间空格，行或列升与处理），"信标"芯6，以及位于该TMA基质3外指定取向芯。虽然这些方法工作以及组织芯片，​​最消耗宝贵的核心TMA空间和组织芯片与差距和空间作为标识符可能会造成混乱的组织核心耗尽了时间。此外，这些方法不适合于在多组织块使用未经阵列形式，因为它们依赖于均匀微阵列芯作为标识符的紧密有序图案异常。大多数非排列多组织块必须适应非均匀组织，根据定义，不显示所述结构化阵列网格使这些象差立场标。 虽然有许多优点，使用一个TMA，最大缺点之一是芯的小尺寸，这并不总是代表很大程度上异质组织1。非排列的多块组织提供了许多的t他好处TMA的，但包含较大的组织样本，或从动物研究整个器官。使用单核组织微阵列具有不同的一致性与基于感兴趣蛋白质整个组织切片和许多需要多个内核以提高一致性7-11。由于复杂性和一些生物标志物的表型异质性，的TMA使用甚至大量芯（&gt; 10）仍然是不充分，并且可能需要的方法比微阵列分析8等。此外，TMA建设费时，技术要求高，需要初始投资成本与投资或访问一个组织阵列。非阵列多的组织块可在任何基本的实验室有显著更少的时间和精力，是需要更多的组织比阵列允许或作为一种手段来削减成本和简化的分析研究，一个有效的替代。 非排列的多块组织同样需要一个可靠的或ientation标记来跟踪样品标识，然而，这些标志物的开发已受到了限制。多描述组织取向的文献集中在正确生理取向为嵌入个人组织片，如纹身的组织与墨12，在处理前13预嵌入琼脂明胶组织，和标记的某些组织与槽口14或缝线15 。虽然功能性的，这些方法是不理想的因为在多组织块标记由于其局限性。缝线将通过快速切片，可能不会在每个部分可见。预嵌入用琼脂明胶可以保持在适当的取向的组织处理和嵌入期间技术，但不提供视觉提示在石蜡块和幻灯片多个样品之间进行区分。在组织上凹口或染料可以复杂化分析或阻塞重要形态细节。另外，组织认同在非排列多组织块可以通过组织片段中的非对称布置嵌入保持，但是这需要3个或更多的组织碎片和可能不允许组织用于分析的最佳安排。 试样方向标记（点）已发展为一种简单，廉价的方法清楚地标识在多组织块组织，并提供比现有方法的方向很多好处。现场是一个小的，丰富多彩的，核心组成羟乙基琼脂糖凝胶处理和组织标记染料，并渗透石蜡（ 图2C）。现货内核嵌入在结束旁边的一个单一的组织在多组织石蜡块，并显示为块，并在每一个部分鲜艳的点（ 图1A - D），清楚地表明了块部分的正确方向为便于组织鉴定。

<table><tbody><tr><td>Histogel Specimen Processing Gel</td><td>Thermo Scientific</td><td>HG-4000-012</td><td>&lt;a href=&quot;http://www.thermoscientific.com/en/product/richard-allan-scientific-histogel-specimen-processing-gel.html&quot; target=&quot;_blank&quot;&gt;http://www.thermoscientific.com/en/product/richard-allan-scientific-histogel-specimen-processing-gel.html&lt;/a&gt;</td></tr><tr><td>Tissue Marking Dye</td><td>Triangle Biomedical Sciences, Inc.</td><td>TMD-5</td><td>Any tissue marking dye would most likely be sufficient.</td></tr><tr><td>Arraymold Kit A 2 mm (60 core)</td><td>Arraymold</td><td>20015A</td><td>Any manual tissue arrayer would work similarly.</td></tr></tbody></table>

spot the correct tissue every time in multi-tissue blocks

多组织石蜡块提供高通量分析以更高的效率，实验的均匀性，并且减少了时间和成本。组织芯片弥补大多数多组织石蜡块，但越来越多的研究人员正在使用含多个人，可以提供很多的组织芯片的优点，但没有规划和设备的大量投资较大的组织非摆着块。任何多组织分析的关键部件是用于识别每个单独的组织的取向方法。虽然方法存在以维持适当的取向，并识别在多组织块的组织，大多不是非常适合于非排列的块，可在阵列内消耗宝贵的空间和/或难以生产在标准组织学实验室。试件方向标签（SPOT）是一种简单，低成本的定向工具，这显然是在石蜡块和所有的组织切片可见FO- [R可靠的标本鉴定阵列和非阵列布局。当场提供了超过现有的取向方法用于非排列块的优点，因为它不需要任何直接修改到组织，并允许在组织片的布置的灵活性。

当场显示为圆形，在石蜡块色彩鲜艳点（ 图1A和图2D），在所有的石蜡切片，并通过H＆E或IHC染色程序保留在玻璃载片上（ 图1B - 1D和2D）。这种明显的视觉提示助剂既在识别每个单独的组织片的组织学技术人员和研究人员，并简化通信作为组织学技术人员可以使用该视觉提示来指示组织片的研究者从幻灯片在显微镜收集数据的排列。组织取向图应包括与提供给研究者的幻灯片和详细解释所以会有在显微分析没有混淆。 图2A示出切片凝固，着色的琼脂糖凝胶塞的结果。 图2B示出嵌入的结果在切片琼脂糖凝胶塞S在石蜡块中。 图2C示出了使用真皮冲，以产生光点核的结果。 <img alt="图1" src="/files/ftp_upload/52868/52868fig1highres.jpg" width="700" /> 图1:现货芯很容易创建和组织块和幻灯片容易看到（A）点显然是在嵌入式模块可见。 （B）的斑点允许容易取向和组织鉴定含有来自不同个体/治疗组出现类似组织的多组织块。 （C）的斑点允许将要用于有价值的组织样品的组织微阵列的每一个核心。现货（D）显微镜查看邻近TMA核心。箭头指示的地方。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52868/52868fig1highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a> </P&gt; <img alt="图2" src="/files/ftp_upload/52868/52868fig2highres.jpg" width="700" /> 图2:斑核的制备 （A） 的凝固羟乙基琼脂糖塞释放从Eppendorf管中并切成横向成5mm切片。 （B）的多片被嵌入在单个石蜡块和（C）一种皮肤冲头用来除去斑点芯。 （ 四）点嵌入到组织芯片的边缘。 <a href="https://www-jove-com.bdigitaluss.remotexs.co/files/ftp_upload/52868/52868fig2highres.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about SpOT the Correct Tissue Every Time in Multi-tissue Blocks at JoVE.com

SpOT the Correct Tissue Every Time in Multi-tissue Blocks

试件定位标记（点）的目的是充当定向工具在单个组织识别在多组织石蜡块中提供帮助。这些协议表明它是如何容易地从普通的，低成本的组织学材料构成，并作为在石蜡块和切片的可靠视觉标记。

现货正确的组织多组织块每一次

Multi-tissue paraffin blocks provide high throughput analysis with increased efficiency, experimental uniformity, and reduced time and cost. Tissue ...

spot-the-correct-tissue-every-time-in-multi-tissue-blocks

Universidad San Sebastian

Research

JoVE Journal

Biology

11.7K 次观看.  Frederick National Laboratory for Cancer Research. 试件定位标记（点）的目的是充当定向工具在单个组织识别在多组织石蜡块中提供帮助。这些协议表明它是如何容易地从普通的，低成本的组织学材料构成，并作为在石蜡块和切片的可靠视觉标记。 

视频: SpOT the Correct Tissue Every Time in Multi-tissue Blocks

Quantitative Multispectral Analysis Following Fluorescent Tissue Transplant for Visualization of Cell Origins, Types, and Interactions

With the desire to understand the contributions of multiple cellular elements to the development of a complex tissue; such as the numerous cell types that participate in regenerating tissue, tumor formation, or vasculogenesis, we devised a multi-colored cellular transplant model of tumor development in which cell populations originate from different fluorescently colored reporter gene mice and are transplanted, engrafted or injected in and around a developing tumor. These colored cells are then recruited and incorporated into the tumor stroma. In order to quantitatively assess bone marrow derived tumor stromal cells, we transplanted GFP expressing transgenic whole bone marrow into lethally irradiated RFP expressing mice as approved by IACUC. 0ovarian tumors that were orthotopically injected into the transplanted mice were excised 6-8 weeks post engraftment and analyzed for bone marrow marker of origin (GFP) as well as antibody markers to detect tumor associated stroma using multispectral imaging techniques. We then adapted a methodology we call MIMicc- Multispectral Interrogation of Multiplexed cellular compositions, using multispectral unmixing of fluoroprobes to quantitatively assess which labeled cell came from which starting populations (based on original reporter gene labels), and as our ability to unmix 4, 5, 6 or more spectra per slide increases, we&#39;ve added additional immunohistochemistry associated with cell lineages or differentiation to increase precision. Utilizing software to detect co-localized multiplexed-fluorescent signals, tumor stromal populations can be traced, enumerated and characterized based on marker staining.1

Complex tissue masses, from organs to tumors, are composed of various cellular elements. We elucidated the contribution of cellular phenotypes within a tissue utilizing multi-labeled fluorescent transgenic mice in combination with multiparameter immunofluorescent staining followed by spectral unmixing to decipher cell origin as well as cell characteristics based on protein expression.

多光谱定量分析继荧光组织移植的细胞起源，类型和相互作用的可视化

With the desire to understand the contributions of multiple cellular elements to the development of a complex tissue; such as the numerous cell types that ...

科研

医学

Automated Dissection Protocol for Tumor Enrichment in Low Tumor Content Tissues

Tumor enrichment in low tumor content tissues, those below 20% tumor content depending on the method, is required to generate quality data reproducibly with many downstream assays such as next generation sequencing. Automated tissue dissection is a new methodology that automates and improves tumor enrichment in these common, low tumor content tissues by decreasing the user-dependent imprecision of traditional macro-dissection and time, cost, and expertise limitations of laser capture microdissection by using digital image annotation overlay onto unstained slides. Here, digital hematoxylin and eosin (H&#38;E) annotations are used to target small tumor areas using a blade that is 250 &#181;m2 in diameter in unstained formalin fixed paraffin embedded (FFPE) or fresh frozen sections up to 20 &#181;m in thickness for automated tumor enrichment prior to nucleic acid extraction and whole exome sequencing (WES). Automated dissection can harvest annotated regions in low tumor content tissues from single or multiple sections for nucleic acid extraction. It also allows for capture of extensive pre- and post-harvest collection metrics while improving accuracy, reproducibility, and increasing throughput with utilization of fewer slides. The described protocol enables digital annotation with automated dissection on animal and/or human FFPE or fresh frozen tissues with low tumor content and could also be used for any region of interest enrichment to boost adequacy for downstream sequencing applications in clinical or research workflows.

Digital annotation with automated tissue dissection provides an innovative approach to enriching tumor in low tumor content cases and is adaptable to both paraffin and frozen tissue types. The described workflow improves accuracy, reproducibility and throughput and could be applied to both research and clinical settings.

用于低肿瘤含量组织中肿瘤富集的自动解剖方案

Tumor enrichment in low tumor content tissues, those below 20% tumor content depending on the method, is required to generate quality data reproducibly ...

癌症研究

Manual Construction of a Tissue Microarray using the Tape Method and a Handheld Microarrayer

The tissue microarray (TMA) is an important research tool in which many formalin fixed paraffin embedded (FFPE) samples can be represented in a single paraffin block. This is achieved by using tissue cores extracted from the region of interest of different donor FFPE blocks and arranging them into a single TMA paraffin block. Once constructed, sections from the completed TMA can be used to perform immunohistochemistry, chromogenic, fluorescence in situ hybridization (FISH) and RNA ISH studies to assess protein expression as well as genomic and transcriptional alterations in many samples simultaneously, thus minimizing tissue usage and reducing reagent costs. There are several different TMA construction techniques. One of the most common construction methods is the recipient method, which works best with cores of the same length for which a minimum length of 4 mm is recommended. Unfortunately, tissue blocks can be heavily resected during the diagnostic process, frequently resulting in "non-ideal" donor block thicknesses of less than 4 mm. The current article and video focus on the double-sided adhesive tape method; an alternative manual, low cost, easy to use, and rapid method to construct low density (&lt;50 cores) TMAs that is highly compatible with these non-ideal donor blocks. This protocol provides a step-by-step guide on how to construct a TMA using this method, with a focus on the critical importance of pathological review and post construction validation.

This protocol outlines the tape method on how to manually construct a tissue microarray using FFPE donor blocks of differing depths.

使用胶带法和手持式微阵列仪手动构建组织微阵列

The tissue microarray (TMA) is an important research tool in which many formalin fixed paraffin embedded (FFPE) samples can be represented in a single ...

绘制肿瘤免疫景观图谱：深入的多重 IHC 分析方案

Multiplex Immunohistochemical Analysis of the Spatial Immune Cell Landscape of the Tumor Microenvironment

The immune cell landscape of the tumor microenvironment potentially contains information for the discovery of prognostic and predictive biomarkers. Multiplex immunohistochemistry is a valuable tool to visualize and identify different types of immune cells in tumor tissues while retaining its spatial information. Here we provide detailed protocols to analyze lymphocyte, myeloid, and dendritic cell populations in tissue sections. Starting from cutting formalin-fixed paraffin-embedded sections, automatic multiplex staining procedures on an automated platform, scanning of the slides on a multispectral imaging microscope, to the analysis of images using an in-house-developed machine learning algorithm ImmuNet. These protocols can be applied to a variety of tumor specimens by simply switching tumor markers to analyze immune cells in different compartments of the sample (tumor versus invasive margin) and apply nearest-neighbor analysis. This analysis is not limited to tumor samples but can also be applied to other (non-)pathogenic tissues. Improvements to the equipment and workflow over the past few years have significantly shortened throughput times, which facilitates the future application of this procedure in the diagnostic setting.

作者聚焦：解锁对肿瘤免疫细胞景观的见解

This protocol describes in detail how immune cell characterization of the tumor microenvironment using multiplex immunohistochemistry is carried out.

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

The immune cell landscape of the tumor microenvironment potentially contains information for the discovery of prognostic and predictive biomarkers. ...

使用 mIHC 破译肺鳞状癌中的肿瘤微环境

Multiplex Immunohistochemistry Staining for Paraffin-embedded Lung Cancer Tissue

Lung cancer is the leading cause of malignant tumor-related morbidity and mortality all over the world, and the complex tumor microenvironment has been considered the leading cause of death in lung cancer patients. The complexity of the tumor microenvironment requires effective methods to understand cell-to-cell relationships in tumor tissues. The multiplex immunohistochemistry (mIHC) technique has become a key tool for inferring the relationship between the expression of proteins upstream and downstream of signaling pathways in tumor tissues and developing clinical diagnoses and treatment plans. mIHC is a multi-label immunofluorescence staining method based on Tyramine Signal Amplification (TSA) technology, which can simultaneously detect multiple target molecules on the same tissue section sample to achieve different protein co-expression and co-localization analysis. In this experimental protocol, paraffin-embedded tissue sections of lung squamous carcinoma of clinical origin were subjected to multiplex immunohistochemical staining. By optimizing the experimental protocol, multiplex immunohistochemical staining of labeled target cells and proteins was achieved, solving the problem of autofluorescence and channel crosstalk in lung tissues. In addition, multiplex immunohistochemical staining is widely used in the experimental validation of tumor-related, high-throughput sequencing, including single-cell sequencing, proteomics, and tissue space sequencing, providing intuitive and visual pathology validation results.

作者聚焦：增强肺癌样本中的多色荧光定位

This experimental protocol describes and optimizes a multiplex immunohistochemistry (IHC) staining method, mainly by optimizing single-channel antibody incubation conditions and adjusting the settings of antibodies and channels to solve the problems of autofluorescence and channel crosstalk in lung cancer tissues of clinical origin.

石蜡包埋肺癌组织的多重免疫组化染色

Lung cancer is the leading cause of malignant tumor-related morbidity and mortality all over the world, and the complex tumor microenvironment has been ...

Production of Tissue Microarrays, Immunohistochemistry Staining and Digitalization Within the Human Protein Atlas

The tissue microarray (TMA) technology provides the means for high-throughput analysis of multiple tissues and cells. The technique is used within the Human Protein Atlas project for global analysis of protein expression patterns in normal human tissues, cancer and cell lines. Here we present the assembly of 1 mm cores, retrieved from microscopically selected representative tissues, into a single recipient TMA block. The number and size of cores in a TMA block can be varied from approximately forty 2 mm cores to hundreds of 0.6 mm cores. The advantage of using TMA technology is that large amount of data can rapidly be obtained using a single immunostaining protocol to avoid experimental variability. Importantly, only limited amount of scarce tissue is needed, which allows for the analysis of large patient cohorts 1 2. Approximately 250 consecutive sections (4 &mu;m thick) can be cut from a TMA block and used for immunohistochemical staining to determine specific protein expression patterns for 250 different antibodies. In the Human Protein Atlas project, antibodies are generated towards all human proteins and used to acquire corresponding protein profiles in both normal human tissues from 144 individuals and cancer tissues from 216 different patients, representing the 20 most common forms of human cancer. Immunohistochemically stained TMA sections on glass slides are scanned to create high-resolution images from which pathologists can interpret and annotate the outcome of immunohistochemistry. Images together with corresponding pathology-based annotation data are made publically available for the research community through the Human Protein Atlas portal (<a href="http://www.proteinatlas.org" target="_blank">www.proteinatlas.org</a>) (Figure 1) 3 4. The Human Protein Atlas provides a map showing the distribution and relative abundance of proteins in the human body. The current version contains over 11 million images with protein expression data for 12.238 unique proteins, corresponding to more than 61% of all proteins encoded by the human genome.

Tissue microarrays allows for an efficient method to gain concurrent information from a multitude of tissues. Representative parts of tissues are assembled into a single paraffin block. Sections from the block are used for immunohistochemistry and analysis of protein expression patterns. Digital scanning generates corresponding images for distribution of data.

组织芯片，​​免疫组织化学染色和数字化生产在人类蛋白质图集

The  tissue microarray (TMA) technology provides the means for high-throughput  analysis of multiple tissues and cells. The technique is used within the ...

现货正确的组织多组织块每一次

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此视频中的章节

多光谱定量分析继荧光组织移植的细胞起源，类型和相互作用的可视化

用于低肿瘤含量组织中肿瘤富集的自动解剖方案

使用胶带法和手持式微阵列仪手动构建组织微阵列

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

石蜡包埋肺癌组织的多重免疫组化染色

石蜡包埋肺癌组织的多重免疫组化染色

石蜡包埋肺癌组织的多重免疫组化染色

组织芯片，免疫组织化学染色和数字化生产在人类蛋白质图集

现货正确的组织多组织块每一次

摘要

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此视频中的章节

多光谱定量分析继荧光组织移植的细胞起源，类型和相互作用的可视化

用于低肿瘤含量组织中肿瘤富集的自动解剖方案

使用胶带法和手持式微阵列仪手动构建组织微阵列

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

肿瘤微环境空间免疫细胞景观的多重免疫组织化学分析

石蜡包埋肺癌组织的多重免疫组化染色

石蜡包埋肺癌组织的多重免疫组化染色

石蜡包埋肺癌组织的多重免疫组化染色

组织芯片，​​免疫组织化学染色和数字化生产在人类蛋白质图集

组织芯片，免疫组织化学染色和数字化生产在人类蛋白质图集