Name: multiplexed fluorescent immunohistochemical staining of four endometrial immune cell types in recurrent miscarriage
Uploaded: 2021-08-04T15:00:00
Description: Watch this Scientific Journal Video about Multiplexed Fluorescent Immunohistochemical Staining of Four Endometrial Immune Cell Types in Recurrent Miscarriage at JoVE.com

本研究于2018年获香港妇产科信托基金及香港健康及医学研究基金（06170186，07180226）资助。

该研究获香港中文大学-新界东区临床研究伦理委员会联合批准。在收集子宫内膜活检之前，从参与者那里获得知情同意。有关控制和 RM 组的包含标准，请参阅简介部分。
1. 样品制备
<ol><li>确保本研究中的所有女性从月经周期的第9天开始每天接受尿液试纸测试，以确定黄体生成素（LH）激增以检测排卵，并在LH激增（LH + 7）后的第 7天精确地进行子宫内膜活检。</li>
	...

<ol>
	<li>ESHRE Guideline Group on RPL et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=ESHRE+guideline:+recurrent+pregnancy+loss.">ESHRE guideline: recurrent pregnancy loss.</a> Human Reproduction Open. 2018 (2), 004 (2018).</li><li>Stirrat, G. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recurrent+miscarriage.">Recurrent miscarriage.</a> Lancet. 336 (8716), 673-675 (1990).</li><li>Rai, R., Regan, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recurrent+miscarriage.">Recurrent miscarriage.</a> Lancet. 368 (9535), 601-611 (2006).</li><li>Royal College of Obstetricians &amp; Gynaecologists. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+investigation+and+treatment+of+couples+with+recurrent+first-trimester+and+second-trimester+miscarriage.+Green-top+Guideline+No.+17.">The investigation and treatment of couples with recurrent first-trimester and second-trimester miscarriage. Green-top Guideline No. 17.</a> Royal College of Obstetricians &amp; Gynaecologists. , (2011).</li><li>King, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Uterine+leukocytes+and+decidualization.">Uterine leukocytes and decidualization.</a> Human Reproduction Update. 6 (1), 28-36 (2000).</li><li>Le Bouteiller, P., Piccinni, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Human+NK+cells+in+pregnant+uterus:+why+there.">Human NK cells in pregnant uterus: why there.</a> American Journal of Reproductive Immunology. 59 (5), 401-406 (2008).</li><li>Lash, G. E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Standardisation+of+uterine+natural+killer+(uNK)+cell+measurements+in+the+endometrium+of+women+with+recurrent+reproductive+failure.">Standardisation of uterine natural killer (uNK) cell measurements in the endometrium of women with recurrent reproductive failure.</a> Journal of Reproductive Immunology. 116, 50-59 (2016).</li><li>Yang, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=HOXA-10+and+E-cadherin+expression+in+the+endometrium+of+women+with+recurrent+implantation+failure+and+recurrent+miscarriage.">HOXA-10 and E-cadherin expression in the endometrium of women with recurrent implantation failure and recurrent miscarriage.</a> Fertility and Sterility. 107 (1), 136-143 (2017).</li><li>Tuckerman, E., Laird, S. M., Prakash, A., Li, T. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Prognostic+value+of+the+measurement+of+uterine+natural+killer+cells+in+the+endometrium+of+women+with+recurrent+miscarriage.">Prognostic value of the measurement of uterine natural killer cells in the endometrium of women with recurrent miscarriage.</a> Human Reproduction. 22 (8), 2208-2213 (2007).</li><li>Jasper, M. J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Macrophage-derived+LIF+and+IL1B+regulate+alpha(1,2)fucosyltransferase+2+(Fut2)+expression+in+mouse+uterine+epithelial+cells+during+early+pregnancy.">Macrophage-derived LIF and IL1B regulate alpha(1,2)fucosyltransferase 2 (Fut2) expression in mouse uterine epithelial cells during early pregnancy.</a> Biology of Reproduction. 84 (1), 179-188 (2011).</li><li>Kopcow, H. D., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=T+cell+apoptosis+at+the+maternal-fetal+interface+in+early+human+pregnancy,+involvement+of+galectin-1.">T cell apoptosis at the maternal-fetal interface in early human pregnancy, involvement of galectin-1.</a> Proceedings of the National Academy of Sciences of the United States of America. 105 (47), 18472-18477 (2008).</li><li>Johnson, P. M., Christmas, S. E., Vince, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Immunological+aspects+of+implantation+and+implantation+failure.">Immunological aspects of implantation and implantation failure.</a> Human Reproduction. 14, 26-36 (1999).</li><li>Hong, G., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Multiplexed+fluorescent+immunohistochemical+staining,+imaging,+and+analysis+in+histological+samples+of+lymphoma.">Multiplexed fluorescent immunohistochemical staining, imaging, and analysis in histological samples of lymphoma.</a> Journal of Visualized Experiments: JoVE. (143), e58711 (2019).</li><li>Carstens, J. L., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spatial+computation+of+intratumoral+T+cells+correlates+with+survival+of+patients+with+pancreatic+cancer.">Spatial computation of intratumoral T cells correlates with survival of patients with pancreatic cancer.</a> Nature Communications. 8, 15095 (2017).</li><li>Zhao, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+use+of+multiplex+staining+to+measure+the+density+and+clustering+of+four+endometrial+immune+cells+around+the+implantation+period+in+women+with+recurrent+miscarriage:+comparison+with+fertile+controls.">The use of multiplex staining to measure the density and clustering of four endometrial immune cells around the implantation period in women with recurrent miscarriage: comparison with fertile controls.</a> Journal of Molecular Histology. 51 (5), 593-603 (2020).</li></ol>

协议中的关键步骤 重要的是要注意，多重染色需要勤奋的优化。使用柠檬酸盐缓冲液和微波技术的抗原检索需要优化，以确保完全剥离抗体并保持组织活力。由于TSA试剂共价结合到抗原周围的位点，它们可以通过空间位阻（也称为"伞效应"）潜在地抑制后续一抗的结合。当多个免疫标志物驻留在单个细胞室中并引起荧光团干扰时，往往会发生这种情况。为了确定是否会有效果，事?...

复发性流产（RM）可定义为妊娠24周前两次或两次以上妊娠的损失1。这种频繁的生殖状况影响了全球高达1%的夫妇2，3。病理生理学是多因素的，可分为胚胎学驱动的原因（主要是由于胚胎核型异常）和影响子宫内膜和/或胎盘发育的母体驱动的原因。这种表现可由父母遗传异常，子宫异常，血栓前疾病，内分泌因素和免疫性疾病引起4。
近年来，免疫效应细胞功能障碍与早孕丢失的发病机制有关5。这激发了许多研究，以阐明月经周期，植入和早孕期间子宫内膜中免疫细胞的特定群体，并在妊娠早期具有特定作用。在这些免疫细胞中，子宫自然杀伤（uNK）细胞在胚胎植入和怀孕期间起着关键作用，特别是在滋养细胞侵袭和血管生成过程中6。研究表明，RM7，8的女性子宫内膜的uNK细胞密度增加，尽管这一发现与流产风险增加无关9。然而，这刺激了研究评估其他免疫细胞类型（如巨噬细胞，子宫树突状细胞）在RM10，11的女性子宫内膜中的密度。然而，仍然不确定RM女性植入围膜内膜的免疫细胞密度是否有显着改变。
对不确定性的一种可能的解释是，由于植入窗口期间子宫内膜的快速变化，评估子宫内膜免疫细胞密度可能很困难。在24小时的时间范围内，子宫内膜的显着变化改变了免疫细胞密度和细胞因子分泌，在这些结果中引入了变异源12。此外，大多数报告主要依赖于使用单细胞染色（例如，传统的IHC方法），无法检查同一组织切片上的多个标记。虽然流式细胞术可用于检测单个样品中的多个细胞群，但所需的大量细胞和耗时的优化阻碍了该方法的普及和效率。因此，多重IHC染色的最新进展可以通过在同一张载玻片上免疫染色多个标记物来评估多个参数，包括细胞谱系和单个免疫亚群的组织学定位，从而解决这个问题。此外，该技术可以在组织可用性有限的情况下最大化所获得的信息。最终，这项技术可以帮助阐明肥沃女性和RM女性之间子宫内膜免疫细胞相互作用的差异。
从威尔士亲王医院招募了两组妇女，包括生育控制妇女（FC）和不明原因复发性流产妇女（RM）。受精控制被定义为至少有一次活产而没有任何自发性流产史的女性，RM女性被定义为在妊娠20周前有≥2次连续流产史的女性。两组受试者符合以下纳入标准：（a）年龄在20至42岁之间，（b）非吸烟者，（c）月经周期正常（25-35天）和正常的子宫结构，（d）子宫内膜活检前至少3个月不使用任何激素方案，（e）通过子宫内膜输卵管造影没有输卵管积水。此外，所有招募的受试者核型正常，三维超声宫腔造影正常，第2天卵泡刺激素<10 IU / L，黄体中期黄体酮>30 nmol /L，甲状腺功能正常，狼疮抗凝剂和抗心磷脂IgG和IgM抗体检测呈阴性。
为了更好地了解RM的免疫学基础，最理想的方法是同时定量和定位植入时子宫内膜中存在的主要免疫细胞类型。本文描述了从样品制备，免疫细胞亚型标记物的多重优化以及载玻片扫描的整个方案，然后进行数据分析，并特别参考检测子宫内膜免疫细胞。此外，本文还描述了如何同时确定子宫内膜中免疫细胞类型的密度和聚类。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Amplification Diluent</td>
			<td>Perkin Elmer</td>
			<td>FP1498</td>
			<td>Fluorophore diluent buffer</td>
		</tr>
		<tr>
			<td>Antibody diluent</td>
			<td>Perkin Elmer</td>
			<td>ARD1001EA</td>
			<td>Diluting the antibody</td>
		</tr>
		<tr>
			<td>CD3</td>
			<td>Spring Bioscience</td>
			<td>M3072</td>
			<td>Primary antibody</td>
		</tr>
		<tr>
			<td>CD20</td>
			<td>Biocare Medical</td>
			<td>CM004B</td>
			<td>Primary antibody</td>
		</tr>
		<tr>
			<td>CD56</td>
			<td>Leica</td>
			<td>NCL-CD56-504</td>
			<td>Primary antibody</td>
		</tr>
		<tr>
			<td>CD68</td>
			<td>Spring Bioscience</td>
			<td>M5510</td>
			<td>Primary antibody</td>
		</tr>
		<tr>
			<td>Citrate Buffer Solution, pH 6.0 (10x)</td>
			<td>Abcam</td>
			<td>AB64214</td>
			<td>Antigen retrieval solution</td>
		</tr>
		<tr>
			<td>EMSURE Xylene (isomeric mixture)</td>
			<td>Merck</td>
			<td>108297</td>
			<td>Dewaxing</td>
		</tr>
		<tr>
			<td>Ethanol absolute</td>
			<td>Merck</td>
			<td>107017</td>
			<td>Ethyl alcohol for rehydration</td>
		</tr>
		<tr>
			<td>HistoCore BIOCUT Manual Rotary Leica Microtome</td>
			<td>Leica</td>
			<td>RM2125RTS</td>
			<td>Sectioning of paraffin-embedded tissue</td>
		</tr>
		<tr>
			<td>inForm Advanced Image Analysis Software</td>
			<td>Perkin Elmer</td>
			<td>inForm&#174; Tissue Finder Software 2.2.1 (version 14.0)</td>
			<td>Data Analysis software</td>
		</tr>
		<tr>
			<td>Mantra&#174; Workstation</td>
			<td>Akoya Biosciences</td>
			<td>CLS140089</td>
			<td>Spectral imaging</td>
		</tr>
		<tr>
			<td>Microwave</td>
			<td>Panasonic</td>
			<td>Inverter</td>
			<td>Microwave stripping</td>
		</tr>
		<tr>
			<td>Opal 520</td>
			<td>Perkin Elmer</td>
			<td>FP1487A</td>
			<td>Appropriate tyramide based fluorescent reagent</td>
		</tr>
		<tr>
			<td>Opal 620</td>
			<td>Perkin Elmer</td>
			<td>FP1495A</td>
			<td>Appropriate tyramide based fluorescent reagent</td>
		</tr>
		<tr>
			<td>Opal 650</td>
			<td>Perkin Elmer</td>
			<td>FP1496A</td>
			<td>Appropriate tyramide based fluorescent reagent</td>
		</tr>
		<tr>
			<td>Opal 690</td>
			<td>Perkin Elmer</td>
			<td>FP1497A</td>
			<td>Appropriate tyramide based fluorescent reagent</td>
		</tr>
		<tr>
			<td>Oven</td>
			<td>Memmert</td>
			<td>U10</td>
			<td>Dewaxing</td>
		</tr>
		<tr>
			<td>Peroxidase Blocking Solution</td>
			<td>DAKO</td>
			<td>S2023</td>
			<td>Removal of tissue peroxidase activities</td>
		</tr>
		<tr>
			<td>Poly-L-lysine coated slide</td>
			<td>FISHER SCIENTIFIC</td>
			<td>120-550-15</td>
			<td>Slide for routine histological use</td>
		</tr>
		<tr>
			<td>PolyHRP Broad Spectrum</td>
			<td>Perkin Elmer</td>
			<td>ARH1001EA</td>
			<td>Secondary antibody</td>
		</tr>
		<tr>
			<td>ProLong&#8482; Gold Antifade Mountant</td>
			<td>ThemoFisher Scientific</td>
			<td>P36930</td>
			<td>Mounting</td>
		</tr>
		<tr>
			<td>Spatstat</td>
			<td>/</td>
			<td>Version 2.1-0</td>
			<td>Spatial point pattern analysis</td>
		</tr>
		<tr>
			<td>Spectral DAPI</td>
			<td>Perkin Elmer</td>
			<td>FP1490A</td>
			<td>Nucleic acid staining</td>
		</tr>
		<tr>
			<td>Tissue Processor</td>
			<td>Thermo Fischer</td>
			<td>Excelsior ES</td>
			<td>Tissue processing for dehydration and paraffination</td>
		</tr>
		<tr>
			<td>Tris Buffer Saline (TBS), 10x</td>
			<td>Cell Signaling Technology</td>
			<td>12498S</td>
			<td>Washing solution</td>
		</tr>
		<tr>
			<td>Tween 20</td>
			<td>Sigma-Aldrich</td>
			<td>P1370-1L</td>
			<td>Nonionic detergent</td>
		</tr>
	</tbody>
</table>

multiplexed fluorescent immunohistochemical staining of four endometrial immune cell types in recurrent miscarriage

免疫组化是生物学研究和临床诊断中组织抗原鉴定和可视化的最常用方法。它可用于表征各种生物过程或病理学，例如伤口愈合，免疫反应，组织排斥和组织 - 生物材料相互作用。然而，使用常规免疫组织化学（IHC）染色在单个组织切片中对多种抗原（特别是对于免疫细胞）的可视化和定量仍然不令人满意。因此，近年来引入了多重技术来鉴定单个组织样品或不同组织样品集合中的多种生物标志物。 
这些技术在区分可育妇女和植入期间复发性流产妇女之间子宫内膜内免疫细胞间相互作用的变化方面特别有用。本文描述了多重荧光IHC染色的详细方案，以研究胚胎植入期间精确定时子宫内膜标本中四种主要免疫细胞类型的密度和聚类。该方法包括样品制备，使用免疫细胞亚型标记物进行多重优化，以及扫描载玻片，然后进行数据分析，并特别参考检测子宫内膜免疫细胞。 
使用这种方法，可以在单个组织切片中同时分析子宫内膜中四种主要免疫细胞类型的密度和聚类。此外，本文还将讨论关键因素和故障排除，以克服所应用的荧光探针之间可能存在的荧光团干扰。重要的是，这种多重染色技术的结果可以帮助深入了解胚胎植入过程中的免疫相互作用和调节。

执行4色多重检测以检测4种子宫内膜免疫细胞类型的总体示意图如图1所示。简而言之，这种多重免疫荧光染色的方案需要8个关键步骤：1.载玻片制备，2.表位检索，3.阻断，4.一抗应用，5.二抗应用，6.信号扩增，7.抗体去除，8.复染和安装。然后使用Mantra工作站进行图像渲染和分析，并使用inForm图像分析软件生成的光谱库，用于区分子宫内膜样品中的4种免疫细胞类型（<strong cl...

Watch this Scientific Journal Video about Multiplexed Fluorescent Immunohistochemical Staining of Four Endometrial Immune Cell Types in Recurrent Miscarriage at JoVE.com

Multiplexed Fluorescent Immunohistochemical Staining of Four Endometrial Immune Cell Types in Recurrent Miscarriage

尽管多重免疫组织化学和多光谱成像取得了进展，但同时表征子宫内膜中主要免疫细胞的密度和聚集仍然是一个挑战。本文描述了一种详细的多重染色方案和成像，用于同时定位子宫内膜中四种免疫细胞类型。

复发性流产中4种子宫内膜免疫细胞类型的多重荧光免疫组织化学染色

Immunohistochemistry is the most commonly used method for the identification and visualization of tissue antigens in biological research and clinical ...

目前的技术允许使用在同一物种中培养的抗体来检测多达七个标记物。演示该程序的将是实验室的技术人员Loucia Chan。首先制备与组织样品一起安装的福尔马林固定石蜡嵌入载玻片，通过将载玻片平放在烤箱中，以60摄氏度烘烤，组织朝上。一小时后，每10分钟用一系列化学物质对载玻片进行脱蜡和再水化，如手稿中所述。将载玻片浸没在tris缓冲的盐水中，然后放入装有甲醇稀释的甲醛混合物的塑料载玻片盒中，以固定样品。接下来，在去离子水中洗涤载玻片两次，持续两分钟。通过将载玻片架置于pH 6.0下充满柠檬酸缓冲液的耐热盒中，继续进行表位检索。然后将盒子放入微波炉中，首先以100%功率加热载玻片50秒，然后以20%功率加热20分钟，以保持相同的温度。在蒸馏水中冲洗载玻片并用TBST洗涤后，将载玻片浸入含有过氧化阻断溶液的罐子中10分钟。孵育后，用蒸馏水冲洗载玻片并用TBST洗涤，然后使用疏水屏障笔在载玻片上的组织部分周围标记边界。然后用TBST冲洗载玻片，用封闭缓冲液覆盖组织切片，并将载玻片在加湿室中孵育15分钟。要除去封闭试剂，首先用感兴趣的一抗孵育载玻片，然后通过用TBST洗涤载玻片三次来除去一抗，然后用标记的聚合物辣根过氧化物酶孵育，二抗。孵育后，在应用用于TSA工作溶液的蛋白石菌群之前洗涤载玻片，然后用抗原取回缓冲液冲洗载玻片。通过以100%功率在微波炉中以100%功率加热抗原检索缓冲液中的载玻片50秒，然后在微波安全容器中加热20%功率20分钟，执行基于微波的剥离。在室温下冷却样品15分钟后，用蒸馏水和TBST冲洗载玻片，然后用dappy溶液孵育5分钟。然后风干载玻片，然后用适当的安装介质安装。在对载玻片进行成像之前，请在工作站中预设相应的滤镜。捕获至少 10 个字段，以便在 200 倍放大倍率下进行分析。要对单元格进行计数，请单击步骤栏中的"对对象进行计数"按钮，以显示对象计数设置面板。如果组织已被分割，请选择要在其中查找对象的组织类别。然后从自动刻度或固定刻度中选择所需的信号缩放。对于对象分割，请从下拉列表中选择主组件。要将接触其他对象的对象检测为单个对象而不是组合对象，请选中最大大小和像素框，然后选择精细分割。接下来，分别计算免疫细胞中的所有基质细胞，以表示数据为免疫细胞相对于每个捕获图像的基质细胞总数的百分比。稍后，将最终单元格计数报告为所有字段的平均值。使用视图编辑器，检查生成的数据表进行后处理，然后导出计数数据表。在代表性分析中，子宫内膜样品中的四种免疫细胞类型被分化。处理后的图像显示组织分割为上皮和基质隔室。对一名育性对照妇女和一名不明原因复发性流产妇女的子宫内膜活检进行了多重免疫染色，在免疫染色中显示代表CD3，CD56，CD68和CD20的单层。一旦免疫声音被鉴定出来，在细胞因子下进一步染色可以帮助解决它们的相互作用和植入成功的机制。

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SCIENTISTS IN ACTION

Preparation of 2-dGuo-Treated Thymus Organ Cultures

2 dGuo治疗胸腺器官培养的制备

In the thymus, interactions between developing T-cell precursors and stromal cells that include cortical and medullary epithelial cells are known to play ...

科研

生物学

Live Cell Imaging of F-actin Dynamics via Fluorescent Speckle Microscopy (FSM)

Live Cell Imaging of F-actin Dynamics via Fluorescent Speckle Microscopy FSM

F - actin的动力学通过荧光活细胞成像斑点显微镜（密克罗尼西亚）

In this protocol we describe the use of Fluorescent Speckle Microscopy (FSM) to capture high-resolution images of actin dynamics in PtK1 cells. A unique ...

Whole-mount Immunohistochemical Analysis for Embryonic Limb Skin Vasculature: a Model System to Study Vascular Branching Morphogenesis in Embryo

全安装胚胎肢体皮肤血管的免疫组织化学分析：一个模型系统，研究在胚胎血管分支形态发生

Whole-mount immunohistochemical analysis for imaging the entire vasculature is pivotal for understanding the cellular mechanisms of branching ...

Analysis of Single-cell Gene Transcription by RNA Fluorescent In Situ Hybridization (FISH)

Analysis of Single-cell Gene Transcription by RNA Fluorescent In Situ Hybridization FISH

单细胞基因转录的RNA荧光分析<em在原位</em&gt;杂交（FISH）

Adhesion of Plasmodium falciparum infected erythrocytes (IE) to human endothelial receptors during malaria infections is mediated by expression of PfEMP1 ...

Separation of Spermatogenic Cell Types Using STA-PUT Velocity Sedimentation

生精细胞类型使用STA把速度沉降分离

Mammalian spermatogenesis is a complex  differentiation process that occurs in several stages in the seminiferous  tubules of the testes. Currently, there ...

Genetic Barcoding with Fluorescent Proteins for Multiplexed Applications

遗传条形码与荧光蛋白的复用的应用程序

Fluorescent proteins, fluorescent dyes and fluorophores in general have revolutionized the field of molecular cell biology. In particular, the discovery ...

Quantitation of Protein Expression and Co-localization Using Multiplexed Immuno-histochemical Staining and Multispectral Imaging

蛋白表达和共定位使用复用免疫组织化学染色和多光谱成像定量

Immunohistochemistry is a commonly used clinical and research lab detection technique for investigating protein expression and localization within ...

Development of an Insert Co-culture System of Two Cellular Types in the Absence of Cell-Cell Contact

在细胞间接触缺失的两个蜂窝型的插入共培养系统的开发

The role of secreted soluble factors in the modification of cellular responses is a recurrent theme in the study of all tissues and systems. In an attempt ...

Isolation and Transcriptome Analysis of Plant Cell Types

植物细胞类型的分离和转录组分析

In multicellular organisms, developmental programming and environmental responses can be highly divergent in different cell types or even within cells, ...

Investigating Zinc Transport Mechanism in Mammalian Cells with FluoZin-3 Assay

Characterizing Mammalian Zinc Transporters Using an In Vitro Zinc Transport Assay

Exploring Cellular Zinc Regulation Through ZnT1 Functionality (Video) | JoVE

使用 体外 锌转运测定表征哺乳动物锌转运蛋白

Transition metals such as Zn2+ ions must be tightly regulated due to their cellular toxicity. Previously, the activity of Zn2+ transporters was measured ...