，NRSA（F32-NS-066698; DPS）;工作是由史密斯家庭基金会（BS），达纳基金会（BS），约翰·默克学者奖励计划（BS），NINDS（BS RO1-NS-07100801）资助项目，南希·劳瑞商标基金会（DPS），美国国立卫生研究院（P30-HD-18655; MRDDRC影像核心）。

1，顺行研资局突触前输入的标签注意:所有涉及使用动物实验，根据美国国立卫生研究院的所有准则审阅及机构动物护理和使用委员会（IACUC）监督。 <ol><li>消毒领域和手段。 </li><li>麻醉用鼠标在一个树脂玻璃感应室4％（体积）的异氟醚（这个体积％异氟醚适用于新生儿，成年小鼠）。观察小鼠的密切合作，以避免过度麻醉。注意:避免吸入用真空废气撤离。 </li><li>经过1-3分钟，（年纪较大的老鼠将需要更少的时间）确保麻醉的适当水平是通过捏尾巴（无反应应观察），并观察呼吸频率（速度应缓慢而稳定的）实现的。 </li><li>把鼠标在其侧面及地方鼻锥递送3-4％（体积）的异氟烷在炉鼻的立体显微镜下（新生儿&lt;产后10天（P10）〜4％（体积）;&gt; P10〜3％（体积））。 </li><li>露出巩膜。为新生儿注射前的眼图张开，用一双小弹簧剪刀剪开眼睑拉背部皮肤，露出巩膜。对于年纪较大的老鼠，用手指拉回来眼睛周围，露出巩膜皮肤。注意:在新生儿中，有时是在用眼角的余光另一个垂直切割是必要的。割双眼皮的角落里时，因为有一个血管，如果切割，会流血过多照顾。 </li><li>使用无菌30.5号针头刺穿一个小孔眼的一侧在巩膜开始处的线。小心避免通过插入针远远不够的斜面进入眼睛损坏镜头。 </li><li>允许玻璃体流出孔，并使用无菌棉签来吸收液体。 </li><li>一旦玻璃已经停止流淌出来的孔，插入连接到汉密尔顿注射器预装了顺行示踪染料的钝端针入孔。注意:插入针缓慢，以避免刺穿眼睛的另一侧或损坏透镜。 </li><li>慢慢的染料注入到眼睛。通常，霍乱毒素β亚基偶联于Alexa的594，647，或488（CTB-594，CTB-647，或CTB-488，5-6微克/微升）用于顺行迹RGC输入。新生儿（≤10页）1-2微升充足且对小鼠&gt; P10〜3微升就足够了。注:Alexa染料是溶酶体降解特别耐</li><li>留针在洞口几秒钟，然后慢慢放开。 </li><li>用棉棒吸去多余的液体，防止染料外泄。 </li><li>取少量抗生素软膏的眼睛。如果眼睛被打开了手术，轻轻地重新定位眼皮在一起。 </li><li>如果注入双眼，在另一只眼睛重复过程。 </li><li>注射后（次），离开鼠标下的热灯或热垫，直到它开始从anesth恢复ESIA。 </li><li>返回鼠标到一个干净的家笼和监控，以确保它是返回到殖民地之前完全清醒。 </li></ol> 2，准备组织成像这个组织制备协议用 ​​于在其中吞噬细胞标记有荧光标记物（ 例如 ，CX3CR1-EGFP为小胶质细胞）报告小鼠。如果记者行不可用，研究者可免疫染色的组织切片（见讨论）。 <ol><li> 〜24小时，在注射后，牺牲了鼠标和解剖大脑。 </li><li>落固定在一个Falcon管中过夜，在4℃下填充有4％多聚甲醛（PFA）的脑注意:PFA是有毒的。通过使用在通风橱中或在下沉气流表和佩戴个人防护装备，避免吸入和皮肤接触。注:固定可短（≥4小时）。另外，代替降定影，4％PFA灌注可用于后跟一个2-24小时滴定位。然而，灌流的比较d来固定掉落新生儿和成人的大脑发现图像质量或定量没有区别。 </li><li>漂洗在磷酸盐缓冲盐水（PBS）中脑3倍。 <ol><li>倒大脑和PFA到一个空的权衡船。 </li><li>用刮刀给大脑转移到另一个称量舟充满PBS。 </li><li>脑洗两次在PBS（转移大脑两个称量皿充满PBS）。 </li></ol></li><li>脑转移到Falcon管中填充了30％的蔗糖溶液。离开大脑中的蔗糖在4℃直至大脑沉到管（24-48小时）的底部。 </li><li>一旦大脑下沉，准备大脑切片。下面的步骤是准备用一个滑动切片机用冷冻阶段40μm的浮动部（低温恒温器也可以被使用）。 <ol><li>用刀片去掉不需要的大脑的部分（外膝体，消除大脑的头端和尾端部分）。 </li><li>冻结胸罩在铝箔在干冰。在这段时间内，冷冻切片机阶段和填充一个24孔板的每个孔用0.5ml的M 0.1的磷酸盐缓冲液（PB）。 </li></ol></li><li>装载冷冻大脑（它应该会出现不透明）在冻结阶段。 <ol><li>取少量的最佳切削温度化合物（OCT）的舞台。 </li><li>一旦华侨城开始冻结，奠定了大脑中的十月将被削减的大脑到底应朝上（ 例如 ，为外膝体，面对尾朝上）。 </li><li>覆盖大脑和OCT具有非常精细粉碎的干冰。 </li><li>离开干冰上脑/华侨城〜30秒。 </li><li>使用大画笔，除去干冰。大脑和OCT应冻结舞台。 </li></ol></li><li>首先通过组织切片，直到感兴趣区域（ROI）的区域为止。 </li><li>一旦投资回报率达到，用小画笔从刀片取出部分，并将它们转移到24孔平台Ë含0.1M PB（步骤2.5.2）。保持油漆刷湿润事先从刀片取出部分润湿它在0.1M的PB。注:部分可留在0.1M PB一夜之间在4°C。 </li><li>一旦部分已收集，可视化的顺行标记荧光解剖显微镜下，选择包含投资回报率的部分。 </li><li>安装在一个用小画笔滑动部分。 <ol><li>适用0.1M的PB一个小型游泳池的充电显微镜幻灯片。 </li><li>转移组织切片在PB的游泳池。 </li><li>用PB和画笔的方向和传播组织。 </li><li>使用的Kimwipe灯芯多余的PB。注意避免排汗关闭部分。 </li><li>允许完全风干。 </li><li>取一小颗的安装介质的每个部分，并安装盖玻片上的顶部（​​22×50毫米，1.5号）。 </li><li>密封滑动的边缘用指甲油和储存在-20°C，直到成像会议。注意:虽然成像在一周的制备忠告，幻灯片可以保持在-20℃下持续数周前成像。 </li></ol></li></ol> 3，组织成像所有图像都在旋转盘共聚焦显微镜（的Ultraview Vox的旋转盘共聚焦显微镜配备二极管激光器（405纳米，445纳米，488纳米，514纳米，561纳米和640纳米））所收购。影像也可以收购任何显微镜获取高分辨率的z栈的能力（ 例如 ，激光扫描共聚焦显微镜，显微镜啶其次是反褶积等）。帧大小通常为1,000×1,000像素。 <ol><li>寻找投资回报率在10倍放大倍率和获取图像。在retinogeniculate系统成像的小胶质细胞，投资回报率是最内侧的外膝体部分。 </li><li>转移到更高的放大倍率（一60X计划阿婆目标，NA = 1.4，通常使用）。 </li><li>收购视图包含使用0.2微米的z一步吞噬细胞第一场秒。 </li><li>获得含15-19每只动物吞噬细胞更多的领域。 </li></ol> 4，准备定量（ImageJ的）图片<ol><li>打开Z堆栈中的ImageJ。 </li><li>转到图像菜单中，选择颜色和分离通道。 </li><li>从含有吞噬物质的通道（s）减去背景。 <ol><li>选择包含吞没材料（ 例如 ，突触前输入）通道窗口。 </li><li>进入处理菜单，并选择减去背景。使用10像素的滚动球半径为于外膝体突触前投入顺行追踪。注:滚球半径凭经验确定。然而，一般应大到不属于背景的部分图像中的最大对象的半径。 </li></ol></li><li>平滑含有吞噬细胞的通道。 <ol><li>选择含有吞噬细胞（ 例如 ，小胶质细胞）的通道窗口。 </li><li>进入处理菜单和SELECT过滤器，平均。使用1.5均值滤波（平滑这在后面的步骤中面绘制的图像）。 </li></ol></li><li>从含有吞噬细胞的通道中减去背景。 <ol><li>选择含有吞噬细胞（ 例如 ，小胶质细胞）的通道窗口。 </li><li>进入处理菜单中，选择减去背景（设置将需要凭经验确定）。使用50像素的EGFP阳性的小胶质细胞滚动球半径。 </li></ol></li><li>通过进入图片菜单，然后选择颜色，合并通道合并通道。 </li><li>作物的ROI含吞噬细胞从合并后的文件（这使得表面渲染速度更快，请参阅步骤5）。 <ol><li>使用绘制矩形选择工具，在工具栏中围绕投​​资回报率一箱。 </li><li>转到图像菜单，然后选择裁剪。 </li></ol></li><li>再次分裂通道（见步骤4.2）。 </li><li>每个通道保存为自己的TIFF文件。 </li></ol> 5，准备用于图像定量在Imaris里<ol><li>打开Imaris里，并确保音量图标在左上角的菜单（ 图2）中选择。 </li><li>裁剪图像的打开第一通道（任何频道都可以先开，只是保持一致）。 </li><li>添加其他通道（s）。进入编辑菜单，选择添加通道和选择文件中的下一个频道。 （重复任何剩余的渠道这一步）。注:要更改颜色，请进入显示调整窗口，点击频道名称（ 图3A）。这将弹出一个对话框来调整颜色。如果显示调整窗口不可见，请进入编辑菜单，然后选择显示调整。 </li><li>调整像素值。进入编辑菜单，然后选择图像属性。在这个窗口中，调整的x，y和z的像素值（这些值取决于显微镜，物镜，摄像头，和z步收购，可以在用于获取图像的软件中找到）。 </li><li>步骤5.5将导致一个非常小图像。要调整，请按一下右下角（ 图2）的拟合图标。 </li><li>在显示调整窗口，调整亮度和对比度，使用左侧和中间的三角形的每个信道。 </li></ol> 6，3维（3D）表面的吞噬细胞的渲染<ol><li>在顶部的工具栏，可以肯定的超越模式的选择（ 图3C）。 </li><li>在显示调整窗口中，取消选中除吞噬细胞通道的所有通道。这会从视野中移除它们。 </li><li>点击面绘制图标（ 图2）。 </li><li>一个创建的窗口将显示在左下角（ 图4A），确保分部的投资回报率是选中在这第一个窗口。点击底部的蓝色前进按钮。 </li><li>调整平滑。 <ol><li>在下一个窗口中，从下拉菜单中（ 图4B）吞噬细胞的通道。这将调整细节量将浮出水面并且应根据经验确定（0.1μm的平滑通常用于小胶质细胞）。 </li><li>选择绝对阈值。 </li><li>点击蓝色的前进按钮在底部。 </li></ol></li><li>阈值的图像。 <ol><li>点击直方图并拖动，直到图像被适当地浮出水面（ 图4C）。这是通过旋转该图像，以确保覆盖在荧光图像上的灰度表面是一个精确的表面表示经验确定。注:要旋转，在指针菜单中选择浏览在上屏幕右侧（ 图3B）中，单击并按住图像旋转。要恢复图像至原来的方向，选择地底从视图菜单中左。 </li><li>点击蓝色的前进按钮在底部。 </li></ol></li><li>在接下来的窗口中，有一个选项按大小等。除非图像是非常嘈杂的过滤掉任何表面，不过滤和删除Ÿ过滤器，自动出现。点击绿色箭头在底部完成面绘制的吞噬细胞。一组新的选项卡会出现（ 图5） </li><li>如有必要，删除不感兴趣的吞噬细胞的一部分的任何表面。 <ol><li>确保在指针菜单（ 图3B）选中选择选项。 </li><li>单击要删除的任何表面，使它变成黄色。若要删除多个曲面，单击表面上，同时按住控制钮。 </li><li>点击铅笔选项卡下删除。 </li></ol></li><li>选择和合并的吞噬细胞的所有表面形成一个表面上。 <ol><li>点击漏斗（ 即过滤器选项卡， 图5C）。 </li><li>单击添加。 </li><li>选择任何过滤器，直方图拖动到最左边（所有表面应为黄色）。 </li><li>回到铅笔选项卡并单击统一。 </li></ol></li><li>进入图形选项卡（ 图5A </sTRONG&gt;），然后单击详细信息，并选择任何参数来显示。选择卷（ 图5D）。对于显示多个参数，去左下角（ 图2）的扳手图标，然后选择参数进行记录。 </li><li>记录吞噬细胞的体积和文件，然后再继续保存。 </li></ol>吞噬材料7。三维表面渲染<ol><li>创建已吞噬细胞引起的吞噬物质的新的信道。 <ol><li>而吞噬细胞表面被选中，单击铅笔标签。 </li><li>单击面膜所有。 </li><li>在出现的对话框中，选择相应的吞噬材料（ 例如 ，突触前投入顺行追踪; 图5B）的通道。 </li><li>检查是否重复申请通道面膜前，点击OK。 </li><li>一个新的通道会出现在一个只包含已吞噬的物质和里面的显示调整窗口吞噬细胞。 </li></ol></li><li>表面渲染含总吞噬和nonengulfed材料的通道。 <ol><li>取消选中在显示调整窗口的所有通道，除了要浮出水面的通道，并取消对吞噬细胞产生的表面。 </li><li>再次点击面绘制图标（见步骤6.3），并利用所描述的吞噬细胞相同的步骤通道创建一个面（步骤6.4-6.11）。注意事项:一定要选择正确的通道前，平滑和阈值（步骤6.5; 图4B）。记下阈值（见步骤6.6; 图4C）。该值也可以在表面渲染后得到完成（魔杖标签; 图5A）。 </li></ol></li><li>记录总的吞噬和非吞噬物质的体积和文件，然后再继续保存。 </li><li>表面渲染吞噬物质（吞噬细胞内的物质）。 <ol><li>可视化的荧光只吞噬物质的新渠道（见步骤7.2.1）。 </li><li>按照上述相同的步骤（步骤7.2-7.3）。注意事项:一定要选择从下拉菜单中选择正确通道平滑之前（选择步骤7.1中创建新的通道）。在阈值窗口中，手动输入被用于渲染的总吞噬和非吞噬材料（见步骤7.2.2）中设定的阈值。 </li></ol></li><li>记录吞噬物质的量和持续之前保存的文件。 </li></ol> 8，计算视场的总容积<ol><li>任何通道创建一个新的表面。 </li><li>在阈值窗口（步骤6.6），滑动直方图一路向左，使整个场浮出水面。 </li><li>堆焊完成整场并记录量（步骤6.7-6.11）。 </li><li>保存该文件。 </li></ol> 9，计算吞噬物质的量<ol><li>计算的密度使用以下算法总吞噬和非吞噬物质的通道: 场（第8步）的总吞噬和非吞噬材料体积（步骤7.3）/音量。 </li><li>使用下面的算法计算出每个单元％吞没: （吞噬物质的量（步骤7.5）/的吞噬细胞（步骤6.11）的总体积）×100 注意:为了计算变化细胞大小，数据被归一化到吞噬细胞的总体积。如果从步骤9.1数字是高度可变的对面域，它可能需要从步骤9.2中的数据归一化，以计算在9.1。 </li></ol>

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W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Development+of+the+vertebrate+neuromuscular+junction.">Development of the vertebrate neuromuscular junction.</a> Annu Rev Neurosci. 22, 389-442 (1999).</li><li>Hua, J. Y., Smith, S. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neural+activity+and+the+dynamics+of+central+nervous+system+development.">Neural activity and the dynamics of central nervous system development.</a> Nat Neurosci. 7 (4), 327-332 (2004).</li><li>Holtmaat, A., Svoboda, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Experience-dependent+structural+synaptic+plasticity+in+the+mammalian+brain.">Experience-dependent structural synaptic plasticity in the mammalian brain.</a> Nat Rev Neurosci. 10 (9), 647-658 (2009).</li><li>Butz, M., 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=Activity-dependent+structural+plasticity.">Activity-dependent structural plasticity.</a> Brain Res Rev. 60 (2), 287-305 (2009).</li><li>Bruel-Jungerman, 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=Brain+plasticity+mechanisms+and+memory:+a+party+of+four.">Brain plasticity mechanisms and memory: a party of four.</a> Neuroscientist. 13 (5), 492-505 (2007).</li><li>Schafer, D. P., Stevens, B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Synapse+elimination+during+development+and+disease:+immune+molecules+take+centre+stage.">Synapse elimination during development and disease: immune molecules take centre stage.</a> Biochem Soc Trans. 38 (2), 476-481 (2010).</li><li>Alexander, A., 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+complement+system:+an+unexpected+role+in+synaptic+pruning+during+development+and+disease.">The complement system: an unexpected role in synaptic pruning during development and disease.</a> Annu Rev Neurosci. 35, 369-389 (2012).</li><li>Brown, A., Weaver, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+dark+side+of+neuroplasticity.">The dark side of neuroplasticity.</a> Exp Neurol. 235 (1), 133-141 (2012).</li><li>Navarro, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chapter+27:+Neural+plasticity+after+nerve+injury+and+regeneration.">Chapter 27: Neural plasticity after nerve injury and regeneration.</a> Int Rev Neurobiol. 87, 483-505 (2009).</li><li>Tan, A. M., Waxman, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Spinal+cord+injury,+dendritic+spine+remodeling,+and+spinal+memory+mechanisms.">Spinal cord injury, dendritic spine remodeling, and spinal memory mechanisms.</a> Exp Neurol. 235 (1), 142-151 (2012).</li><li>Wolpaw, J. R., Tennissen, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activity-dependent+spinal+cord+plasticity+in+health+and+disease.">Activity-dependent spinal cord plasticity in health and disease.</a> Annu Rev Neurosci. 24, 807-843 (2001).</li><li>Schafer, D. P., 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+&amp;#34;quad-partite&amp;#34;+synapse:+Microglia-synapse+interactions+in+the+developing+and+mature+CNS.">The &amp;#34;quad-partite&amp;#34; synapse: Microglia-synapse interactions in the developing and mature CNS.</a> Glia. 61 (1), 24-36 (2013).</li><li>Mukhopadhyay, S., 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=Manganese-induced+trafficking+and+turnover+of+the+cis-Golgi+glycoprotein+GPP130.">Manganese-induced trafficking and turnover of the cis-Golgi glycoprotein GPP130.</a> Mol Biol Cell. 21 (7), 1282-1292 (2010).</li><li>Jung, S., 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=Analysis+of+fractalkine+receptor+CX(3)CR1+function+by+targeted+deletion+and+green+fluorescent+protein+reporter+gene+insertion.">Analysis of fractalkine receptor CX(3)CR1 function by targeted deletion and green fluorescent protein reporter gene insertion.</a> Mol Cell Biol. 20, 4106-4114 (2000).</li><li>Knobloch, M., Mansuy, I. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dendritic+spine+loss+and+synaptic+alterations+in+Alzheimer's+disease.">Dendritic spine loss and synaptic alterations in Alzheimer's disease.</a> Mol Neurobiol. 37 (1), 73-82 (2008).</li><li>Masliah, 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=Synaptic+remodeling+during+aging+and+in+Alzheimer's+disease.">Synaptic remodeling during aging and in Alzheimer's disease.</a> J Alzheimers Dis. 9 (3 Suppl), 91-99 (2006).</li><li>Yoshiyama, 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=Synapse+loss+and+microglial+activation+precede+tangles+in+a+P301S+tauopathy+mouse+model.">Synapse loss and microglial activation precede tangles in a P301S tauopathy mouse model.</a> Neuron. 53 (3), 337-351 (1016).</li><li>Milnerwood, A. J., Raymond, L. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Early+synaptic+pathophysiology+in+neurodegeneration:+insights+from+Huntington's+disease.">Early synaptic pathophysiology in neurodegeneration: insights from Huntington's disease.</a> Trends Neurosci. 33 (11), 513-523 (2010).</li><li>Noda, M., Suzumura, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sweepers+in+the+CNS:+Microglial+Migration+and+Phagocytosis+in+the+Alzheimer+Disease+Pathogenesis.">Sweepers in the CNS: Microglial Migration and Phagocytosis in the Alzheimer Disease Pathogenesis.</a> Int J Alzheimers Dis. , (2012).</li><li>Rotshenker, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microglia+and+macrophage+activation+and+the+regulation+of+complement-receptor-3+(CR3/MAC-1)-mediated+myelin+phagocytosis+in+injury+and+disease.">Microglia and macrophage activation and the regulation of complement-receptor-3 (CR3/MAC-1)-mediated myelin phagocytosis in injury and disease.</a> J Mol Neurosci. 21 (1), 65-72 (2003).</li><li>Smith, M. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Phagocytosis+of+myelin+in+demyelinative+disease:+a+review.">Phagocytosis of myelin in demyelinative disease: a review.</a> Neurochem Res. 24 (2), 261-268 (1999).</li></ol>

没有利益冲突的声明。

为了准确地测量吞噬作用，吞噬材料必须贴在这样一种方式，研究人员可以直观一次溶酶体降解发生。此外，高分辨率成像是必需的，随后使用软件，使研究者能够可视化整个细胞的体积和量化其内容。在这个协议中，我们描述了用于使用CTB偶联至Alexa染料来标记吞噬材料结合高分辨率共焦显微镜和三维重建测定吞噬细胞介导的​​吞噬一个高度可靠的和定量的方法。这种方法已被用来在实验室?...

<html>突触电路改造整个动物的生命。在脑发育，突触形成过剩，而且必须经过突触的修剪涉及选择性去除突触的一个子集和那些仍然8-10突触的维持和加强。这个过程是必需的，以实现对成人神经系统的精确的连接特性。在成人中，突触也可以是塑料，特别是在学习和记忆的上下文中。此可塑性的结构相互关系被认为是包括树突棘和突触前boutons 11-13的加法和/或消除。除了 ​​在健康的神经系统这些角色，突触重塑也参与了神经系统疾病/伤害12,14,15。例如，下面的脊髓损伤，轴突切断后必须改造，形成新的突触，实现功能的恢复16-19。 NT"&gt;新兴突触可塑性的一个重要方面是吞噬或拟用于去除3,5,20突触吞噬的过程中，我们最近发现这种现象在健康，出生后小鼠大脑突触7修剪的情况下，特别，小胶质细胞，常住中枢神经系统的免疫细胞和吞噬细胞，被证明在高峰期和发育突触的修剪，丘脑的产后背外侧膝状体核（外膝体）的区域吞没突触前输入。这吞没的遗传或药理封锁导致突触连接持续的赤字。 在这个协议中，我们描述了一种可靠的和高度的定量测定法来测量的突触前输入，吞噬细胞介导的​​吞噬。对于这篇文章的目的，此法将在发展中retinogeniculate系统，其中包括视网膜神经节细胞（RGC的）居住在视网膜的背景下提出的项目突触前输入到外膝体（ 图1A）。开始，溶酶体降解抗性顺行标记策略进行说明，这是用于可视化的外膝体RGC特异突触前输入（ 图1）7,21。以下本说明书中，用于成像的详细方法和使用共聚焦显微镜并结合3维（3D）表面体绘制的定量测定吞噬将得到。此方法是基于固定的组织制备，但也可以适于在实时成像研究使用。重要的是，尽管已经测定在健康，产后retinogeniculate系统的上下文中得到证实，人们可以应用相同的技术来评估其它吞噬细胞 - 神经元相互作用的整个大脑和疾病过程中，以及在其他器官系统的吞噬细胞的功能。

<table><tbody><tr><td>Heat pad</td><td>Vet Equip, Inc.</td><td>965500&amp;nbsp;</td><td></td></tr><tr><td>Warm water source for heat pad</td><td>Kent Scientific</td><td>TP-700</td><td></td></tr><tr><td>Stereo microscope</td><td>DSC Optical</td><td>Zeiss Opmi -6 Surgical Microscope</td><td></td></tr><tr><td>Sliding microtome with freezing stage</td><td>Leica</td><td>SM2010 R</td><td></td></tr><tr><td>Microtome blade</td><td>Leica</td><td>14021607100</td><td></td></tr><tr><td>Fluorescent dissecting microscope</td><td>Nikon</td><td>SMZ800 with Epi-fluorescence attachment</td><td></td></tr><tr><td>Spinning disk confocal microscope</td><td>Perkin Elmer</td><td>UltraView Vox Spinning Disk Confocal</td><td></td></tr><tr><td>10 &amp;micro;l Hamilton gas tight syringes</td><td>Hamilton</td><td>80030</td><td>Use a different syringe for each color dye/tracer</td></tr><tr><td>Hamilton needles</td><td>Hamilton</td><td>7803-05, specifications: blunt, 1.5&quot;</td><td></td></tr><tr><td>Alexa-conjugated cholera toxin &amp;beta; subunit (CTB)</td><td>Invitrogen</td><td>488: C22841</td><td>Reconstitute in sterile saline, 80 &amp;micro;l (488), 100 &amp;micro;l (594), 20 &amp;micro;l (647)</td></tr><tr><td>594: C22842</td><td></td><td></td><td></td></tr><tr><td>647: C34778</td><td></td><td></td><td></td></tr><tr><td>Phosphate Buffered Saline&amp;nbsp;(PBS)</td><td>Sigma</td><td>P4417-50TAB</td><td></td></tr><tr><td>Neomycin and Polymyxin B Sulfates and Bacitracin Zinc Ophthalmic Ointment USP (antibiotic ointment)</td><td>Bausch &amp;amp; Lomb</td><td>24208-780-55</td><td></td></tr><tr><td>30.5 G needle</td><td>Becton Dickinson</td><td>305106</td><td></td></tr><tr><td>Spring scissors</td><td>Roboz</td><td>RS-5630</td><td></td></tr><tr><td>Cotton-tipped applicator</td><td>Fisher</td><td>23-400-125</td><td></td></tr><tr><td>Paraformaldeyde (PFA)</td><td>Electron Microscopy Sciences</td><td>15710</td><td>Dilute 16% to 4% in PBS. Paraformaldehye is toxic, use in a fume hood and wear personal protective equipment.</td></tr><tr><td>Dissection tools &amp;ndash; scissors, forceps, spatula</td><td>Small scissors: Fine Science Tools</td><td>Small scissors:14370-22</td><td></td></tr><tr><td>Large scissors: Roboz</td><td>Large scissors: RS-6820</td><td></td><td></td></tr><tr><td>#55 forceps: Fine Science Tools</td><td>#55 forceps: 11255-20</td><td></td><td></td></tr><tr><td>Spatula: Ted Pella, Inc.</td><td>Spatula: 13504</td><td></td><td></td></tr><tr><td>Sucrose</td><td>Sigma</td><td>S8501-5KG</td><td>Make 30% sucrose in PBS (weight/vol)</td></tr><tr><td>OCT Compound</td><td>VWR</td><td>25608-930</td><td></td></tr><tr><td>Weigh boat</td><td>USA Scientific</td><td>2347-1426</td><td></td></tr><tr><td>24-well plates</td><td>BD Biosciences</td><td>353047</td><td></td></tr><tr><td>Sodium phosphate monobasic</td><td>Sigma</td><td>S6566-500G</td><td>Make 0.2 M sodium phosphate monobasic (PB-A) in ddH&lt;sub&gt;2&lt;/sub&gt;O and 0.2 M sodium phosphate dibasic (PB-B) in ddH&lt;sub&gt;2&lt;/sub&gt;O. To make 0.1 M PB, combine 19 ml PB-A and 81 ml PB-B, fill to 200 ml with ddH&lt;sub&gt;2&lt;/sub&gt;O</td></tr><tr><td>Sodium phosphate dibasic&amp;nbsp;</td><td>Sigma</td><td>S5136-500G</td><td></td></tr><tr><td>Coverslips, 22 X 50 mm, No. 1.5</td><td>VWR</td><td>48393 194</td><td></td></tr><tr><td>Charged microscope slide</td><td>VWR</td><td>48311-703</td><td></td></tr><tr><td>Vectasheild</td><td>Vector Laboratories</td><td>H-1200</td><td></td></tr></tbody></table>

an engulfment assay: a protocol to assess interactions between cns phagocytes and neurons

吞噬作用是一个过程，其中一个细胞吞噬材料（整个小区的小区，杂物等部件）在其周围的细胞外环境，并随后消化此材料，通常通过溶酶体降解。小胶质细胞是中枢神经系统（CNS）的细胞的吞噬功能的广泛的神经退行性疾病（ 例如 ，β-淀粉样蛋白的间隙在阿尔茨海默氏病）的健康脑的发展（ 例如，突触条件已经描述的驻地免疫细胞修剪）1-6。下面的协议是制定可视化和量化，在发育中的小鼠retinogeniculate系统7的突触前输入，小神经胶质细胞介导的吞噬的吞噬测定。虽然这种测定法用于评估在此特定情况下的小胶质细胞的功能，类似的方法可用于评估其他吞噬细胞在整个脑（ 例如，星形胶质细胞）和身体的其余部分（ 例如 ，外周血巨噬细胞）以及在其中突触重塑发生（ 例如 ，脑损伤/疾病）其他上下文中。

最近，我们使用这个吞噬试验，以可视化和量化的显影retinogeniculate系统的突触前输入，小神经胶质细胞介导的吞噬（ 图1）7。从CX3CR1-EGFP杂合子小鼠视网膜神经节细胞进行顺行追踪与CTB-594和CTB-647进入左眼和右眼分别。在此之后追踪，外膝体中EGFP阳性的小胶质细胞进行成像。这些图像随后的表面呈现为体积测量。 使用这种技术，我们发现，在外膝体（P5?...

Watch this Scientific Journal Video about An Engulfment Assay: A Protocol to Assess Interactions Between CNS Phagocytes and Neurons at JoVE.com

An Engulfment Assay: A Protocol to Assess Interactions Between CNS Phagocytes and Neurons

小胶质细胞是中枢神经系统（CNS）具有高容量，以在它们的胞外环境吞噬或吞噬物质的驻留的免疫细胞。这里，用于可视化和测量突触小部件的小神经胶质细胞介导的​​吞噬一个广泛适用的，可靠的和高度的定量测定法进行说明。

一个吞噬含量:一个协议，以评估之间的中枢神经系统吞噬细胞和神经元的相互作用

Phagocytosis is a process in which a cell engulfs material (entire cell, parts of a cell, debris, etc.) in its surrounding extracellular environment and ...

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Research

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Neuroscience

18.2K 次观看.  Boston Children's Hospital, Harvard Medical School. 小胶质细胞是中枢神经系统（CNS）具有高容量，以在它们的胞外环境吞噬或吞噬物质的驻留的免疫细胞。这里，用于可视化和测量突触小部件的小神经胶质细胞介导的​​吞噬一个广泛适用的，可靠的和高度的定量测定法进行说明。 

视频: An Engulfment Assay: A Protocol to Assess Interactions Between CNS Phagocytes and Neurons

Detection of MicroRNAs in Microglia by Real-time PCR in Normal CNS and During Neuroinflammation

Microglia are cells of the myeloid lineage that reside in the central nervous system (CNS)1. These cells play an important role in pathologies of many diseases associated with neuroinflammation such as multiple sclerosis (MS)2. Microglia in a normal CNS express macrophage marker CD11b and exhibit a resting phenotype by expressing low levels of activation markers such as CD45. During pathological events in the CNS, microglia become activated as determined by upregulation of CD45 and other markers3. The factors that affect microglia phenotype and functions in the CNS are not well studied. MicroRNAs (miRNAs) are a growing family of conserved molecules (~22 nucleotides long) that are involved in many normal physiological processes such as cell growth and differentiation4 and pathologies such as inflammation5. MiRNAs downregulate the expression of certain target genes by binding complementary sequences of their mRNAs and play an important role in the activation of innate immune cells including macrophages6 and microglia7. In order to investigate miRNA-mediated pathways that define the microglial phenotype, biological function, and to distinguish microglia from other types of macrophages, it is important to quantitatively assess the expression of particular microRNAs in distinct subsets of CNS-resident microglia. Common methods for measuring the expression of miRNAs in the CNS include quantitative PCR from whole neuronal tissue and in situ hybridization. However, quantitative PCR from whole tissue homogenate does not allow the assessment of the expression of miRNA in microglia, which represent only 5-15% of the cells of neuronal tissue. Hybridization in situ allows the assessment of the expression of microRNA in specific cell types in the tissue sections, but this method is not entirely quantitative. In this report we describe a quantitative and sensitive method for the detection of miRNA by real-time PCR in microglia isolated from normal CNS or during neuroinflammation using experimental autoimmune encephalomyelitis (EAE), a mouse model for MS. The described method will be useful to measure the level of expression of microRNAs in microglia in normal CNS or during neuroinflammation associated with various pathologies including MS, stroke, traumatic injury, Alzheimer's disease and brain tumors.

Microglia are resident macrophages that provide the first line of defense and immune surveillance of the central nervous system. MicroRNAs are regulatory molecules that play an important role in many physiological processes including activation and differentiation of macrophages. In this article, we describe the method for measurement of microRNAs in microglia.

小胶质细胞的小分子RNA检测在正常中枢神经系统的实时PCR技术在神经发炎

Microglia are cells of the myeloid lineage that reside in the central nervous system (CNS)1. These cells play an important role in pathologies of many ...

科研

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Assessing Retinal Microglial Phagocytic Function In Vivo Using a Flow Cytometry-based Assay

Microglia are the tissue resident macrophages of the central nervous system (CNS) and they perform a variety of functions that support CNS homeostasis, including phagocytosis of damaged synapses or cells, debris, and/or invading pathogens. Impaired phagocytic function has been implicated in the pathogenesis of diseases such as Alzheimer's and age-related macular degeneration, where amyloid-&#946; plaque and drusen accumulate, respectively. Despite its importance, microglial phagocytosis has been challenging to assess in vivo. Here, we describe a simple, yet robust, technique for precisely monitoring and quantifying the in vivo phagocytic potential of retinal microglia. Previous methods have relied on immunohistochemical staining and imaging techniques. Our method uses flow cytometry to measure microglial uptake of fluorescently labeled particles after intravitreal delivery to the eye in live rodents. This method replaces conventional practices that involve laborious tissue sectioning, immunostaining, and imaging, allowing for more precise quantification of microglia phagocytic function in just under six hours. This procedure can also be adapted to test how various compounds alter microglial phagocytosis in physiological settings. While this technique was developed in the eye, its use is not limited to vision research.

Assessing Retinal Microglial Phagocytic Function In Vivo Using a Flow Cytometry-based Assay

Microglial phagocytosis is critical for the maintenance of tissue homeostasis and inadequate phagocytic function has been implicated in pathology. However, assessing microglia function in vivo is technically challenging. We have developed a simple but robust technique for precisely monitoring and quantifying the phagocytic potential of microglia in a physiological setting.

评估视网膜小胶质细胞吞噬功能<em&gt;在体内</em&gt;使用基于流式细胞术流量测定

Microglia are the tissue resident macrophages of the central nervous system (CNS) and they perform a variety of functions that support CNS homeostasis, ...

A Novel In Vitro Live-imaging Assay of Astrocyte-mediated Phagocytosis Using pH Indicator-conjugated Synaptosomes

Astrocytes are the major cell type in the brain and directly contact synapses and blood vessels. Although microglial cells have been considered the major immune cells and only phagocytes in the brain, recent studies have shown that astrocytes also participate in various phagocytic processes, such as developmental synapse elimination and clearance of amyloid beta plaques in Alzheimer&#39;s disease (AD). Despite these findings, the efficiency of astrocyte engulfment and degradation of their targets is unclear compared with that of microglia. This lack of information is mostly due to the lack of an assay system in which the kinetics of astrocyte- and microglia-mediated phagocytosis are easily comparable. To achieve this goal, we have developed a long-term live-imaging in vitro phagocytosis assay to evaluate the phagocytic capacity of purified astrocytes and microglia. In this assay, real-time detection of engulfment and degradation is possible using pH indicator-conjugated synaptosomes, which emit bright red fluorescence in acidic organelles, such as lysosomes. Our novel assay provides simple and effective detection of phagocytosis through live-imaging. In addition, this in vitro phagocytosis assay can be used as a screening platform to identify chemicals and compounds that can enhance or inhibit the phagocytic capacity of astrocytes. As synaptic pruning malfunction and pathogenic protein accumulation have been shown to cause mental disorders or neurodegenerative diseases, chemicals and compounds that modulate the phagocytic capacity of glial cells should be helpful in treating various neurological disorders.

A Novel In Vitro Live-imaging Assay of Astrocyte-mediated Phagocytosis Using pH Indicator-conjugated Synaptosomes

This protocol presents an in vitro live-imaging phagocytosis assay to measure the phagocytic capacity of astrocytes. Purified rat astrocytes and microglia are used along with pH indicator-conjugated synaptosomes. This method can detect real-time engulfment and degradation kinetics and provides a suitable screening platform to identify factors modulating astrocyte phagocytosis.

一种新的体外活体成像法测定星形胶质细胞的吞噬作用 pH 指标-共轭突

Astrocytes are the major cell type in the brain and directly contact synapses and blood vessels. Although microglial cells have been considered the major ...

神经科学

Experimental Analysis of Apoptotic Thymocyte Engulfment by Macrophages

Cell apoptosis is a natural process and plays a critical role in embryonic development, homeostatic regulation, immune tolerance induction, and resolution of inflammation. Accumulation of apoptotic debris in the body may trigger chronic inflammatory responses that lead to systemic autoimmune diseases over time. Impaired apoptotic cell clearance has been implicated in a variety of autoimmune diseases. Apoptotic clearance is a complex process rarely detected under physiological conditions. It involves abundant surface receptors and signaling molecules. Studying the process of apoptotic cell clearance provides insightful molecular mechanisms and subsequent biological responses, which may lead to the development of new therapeutics. Here, we describe protocols for the induction of apoptotic thymocytes, the preparation of peritoneal macrophages, and the analysis of apoptotic cell clearance by flow cytometry and microscopy. All cells will undergo apoptosis at a certain stage, and many residential and circulating cells can uptake apoptotic debris. Therefore, the protocol described here can be used in many applications to characterize apoptotic cell binding and ingestion by many other cell types.

Here, we present a protocol to prepare apoptotic thymocytes and peritoneal macrophages and analyze the efficiency of efferocytosis and the specific inhibitor-mediated blocking of apoptotic thymocytes engulfment. This protocol has a broad application in cell-mediated clearance of other particles including artificial beads and bacteria.

巨噬细胞吞噬胸腺细胞吞噬的实验分析

Cell apoptosis is a natural process and plays a critical role in embryonic development, homeostatic regulation, immune tolerance induction, and resolution ...

Flow Cytometry Approach to Characterize Phagocytic Properties of Acutely-Isolated Adult Microglia and Brain Macrophages In Vitro

Microglia and central nervous system (CNS)-infiltrating macrophages, collectively called CNS mononuclear phagocytes (CNS-MPs), play central roles in neurological diseases including neurodegeneration and stroke. CNS-MPs are involved in phagocytic clearance of pathological proteins, debris and neuronal synapses, each with distinct underlying molecular pathways. Characterizing these phagocytic properties can provide a functional readout that compliments molecular profiling of microglia using traditional flow cytometry, transcriptomics and proteomics approaches. Phagocytic profiling of microglia has relied on microscopic visualization and in vitro cultures of mouse neonatal microglia. The former approach suffers from limited sampling while the latter approach is inherently poorly reflective of the true in vivo state of adult CNS-MPs. This paper describes optimized protocols to phenotype phagocytic properties of acutely-isolated mouse CNS-MPs by flow cytometry. CNS-MPs are acutely isolated from adult mouse brain using mechanical dissociation followed by density gradient centrifugation, incubated with fluorescent microspheres or fluorescent A&#946; fibrils, washed, and then labeled with panels of antibodies against surface markers (CD11b, CD45). Using this approach, it is possible to compare phagocytic properties of brain-resident microglia with CNS-infiltrating macrophages and then assess the effect of aging and disease pathology on these phagocytic phenotypes. This rapid method also holds potential to functionally phenotype acutely-isolated human CNS-MPs from post-mortem or surgical brain specimens. Additionally, specific mechanisms of phagocytosis by CNS-MP subsets can be investigated by inhibiting select phagocytic pathways.

Assessment of phagocytic properties of microglia and brain macrophages can provide a valuable functional dimension to molecular profiling studies. We describe a validated protocol that uses flow cytometry to rapidly and reliably quantify phagocytosis of fluorescent microspheres and amyloid beta fibrils by acutely-isolated microglia and brain macrophages from mouse models.

Microglia and central nervous system (CNS)-infiltrating macrophages, collectively called CNS mononuclear phagocytes (CNS-MPs), play central roles in ...

In vitro Quantitative Imaging Assay for Phagocytosis of Dead Neuroblastoma Cells by iPSC-Macrophages

Microglia orchestrate neuroimmune responses in several neurodegenerative diseases, including Parkinson&#39;s disease and Alzheimer&#39;s disease. Microglia clear up dead and dying neurons through the process of efferocytosis, a specialized form of phagocytosis. The phagocytosis function can be disrupted by environmental or genetic risk factors that affect microglia. This paper presents a rapid and simple in vitro&#160;microscopy protocol for studying microglial efferocytosis in an induced pluripotent stem cell (iPSC) model of microglia, using a human neuroblastoma cell line (SH-SY5Y) labeled with a pH-sensitive dye for the phagocytic cargo. The procedure results in a high yield of dead neuroblastoma cells, which display surface phosphatidylserine, recognized as an &#34;eat-me&#34; signal by phagocytes. The 96-well plate assay is suitable for live-cell time-lapse imaging, or the plate can be successfully fixed prior to further processing and quantified by high-content microscopy. Fixed-cell high-content microscopy enables the assay to be scaled up for screening of small molecule inhibitors or assessing the phagocytic function of genetic variant iPSC lines. While this assay was developed to study phagocytosis of whole dead neuroblastoma cells by iPSC-macrophages, the assay can be easily adapted for other cargoes relevant to neurodegenerative diseases, such as synaptosomes and myelin, and other phagocytic cell types.

Neurodegenerative diseases are associated with dysregulated microglia functions. This article outlines an in vitro assay of phagocytosis of neuroblastoma cells by iPSC-macrophages. Quantitative microscopy readouts are described for both live-cell time-lapse imaging and fixed-cell high-content imaging.

iPSC-巨噬细胞死亡神经母细胞的体外定量成像检测

Microglia orchestrate neuroimmune responses in several neurodegenerative diseases, including Parkinson&#39;s disease and Alzheimer&#39;s disease. ...

Human Microglia-like Cells: Differentiation from Induced Pluripotent Stem Cells and In Vitro Live-cell Phagocytosis Assay using Human Synaptosomes

Microglia are the resident immune cells of myeloid origin that maintain homeostasis in the brain microenvironment and have become a key player in multiple neurological diseases. Studying human microglia in health and disease represents a challenge due to the extremely limited supply of human cells. Induced pluripotent stem cells (iPSCs) derived from human individuals can be used to circumvent this barrier. Here, it is demonstrated how to differentiate human iPSCs into microglia-like cells (iMGs) for in vitro experimentation. These iMGs exhibit the expected and physiological properties of microglia, including microglia-like morphology, expression of proper markers, and active phagocytosis. Additionally, documentation for isolating and labeling synaptosome substrates derived from human iPSC-derived lower motor neurons (i3LMNs) is provided. A live-cell, longitudinal imaging assay is used to monitor engulfment of human synaptosomes labeled with a pH-sensitive dye, allowing for investigations of iMG's phagocytic capacity. The protocols described herein are broadly applicable to different fields that are investigating human microglia biology and the contribution of microglia to disease.

Human Microglia-like Cells: Differentiation from Induced Pluripotent Stem Cells and In Vitro Live-cell Phagocytosis Assay using Human Synaptosomes

This protocol describes the differentiation process of human induced pluripotent stem cells (iPSCs) into microglia-like cells for in vitro experimentation. We also include a detailed procedure for generating human synaptosomes from iPSC-derived lower motor neurons that can be used as a substrate for in vitro phagocytosis assays using live-cell imaging systems.

人小胶质细胞样细胞:使用人突触体从诱导多能干细胞和 体外 活细胞吞噬测定中分化

Microglia are the resident immune cells of myeloid origin that maintain homeostasis in the brain microenvironment and have become a key player in multiple ...

基于流式细胞术的突触材料小胶质细胞吞噬检测

Quantification of Microglial Engulfment of Synaptic Material Using Flow Cytometry

Microglia play a pivotal role in synaptic refinement in the brain. Analysis of microglial engulfment of synapses is essential for comprehending this process; however, currently available methods for identifying microglial engulfment of synapses, such as immunohistochemistry (IHC) and imaging, are laborious and time-intensive. To address this challenge, herein we present in vitro and in vivo* assays that allow fast and high-throughput quantification of microglial engulfment of synapses using flow cytometry.
In the in vivo* approach, we performed intracellular vGLUT1 staining following fresh cell isolation from adult mouse brains to quantify engulfment of vGLUT1+ synapses by microglia. In the in vitro synaptosome engulfment assay, we used freshly isolated cells from the adult mouse brain to quantify the engulfment of pHrodo&#160;Red-labeled synaptosomes by microglia. These protocols together provide a time-efficient approach to quantifying microglial engulfment of synapses and represent promising alternatives to labor-intensive image analysis-based methods. By streamlining the analysis, these assays can contribute to a better understanding of the role of microglia in synaptic refinement in different disease models.

作者聚焦：通过心理神经免疫学的技术创新促进理解

Here we present two protocols to quantify microglial engulfment of vGLUT1-positive synapses and pHRodo&#160;Red-labeled crude synaptosomes using flow cytometry.

使用流式细胞术定量突触材料的小胶质细胞吞噬

Microglia play a pivotal role in synaptic refinement in the brain. Analysis of microglial engulfment of synapses is essential for comprehending this ...

一个吞噬含量:一个协议，以评估之间的中枢神经系统吞噬细胞和神经元的相互作用

摘要

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此视频中的章节

小胶质细胞的小分子RNA检测在正常中枢神经系统的实时PCR技术在神经发炎

评估视网膜小胶质细胞吞噬功能

一种新的体外活体成像法测定星形胶质细胞的吞噬作用 pH 指标-共轭突

巨噬细胞吞噬胸腺细胞吞噬的实验分析

Flow Cytometry Approach to Characterize Phagocytic Properties of Acutely-Isolated Adult Microglia and Brain Macrophages In Vitro

iPSC-巨噬细胞死亡神经母细胞的体外定量成像检测

人小胶质细胞样细胞:使用人突触体从诱导多能干细胞和体外活细胞吞噬测定中分化

使用流式细胞术定量突触材料的小胶质细胞吞噬

使用流式细胞术定量突触材料的小胶质细胞吞噬

使用流式细胞术定量突触材料的小胶质细胞吞噬

一个吞噬含量:一个协议，以评估之间的中枢神经系统吞噬细胞和神经元的相互作用

摘要

探索更多视频

此视频中的章节

小胶质细胞的小分子RNA检测在正常中枢神经系统的实时PCR技术在神经发炎

评估视网膜小胶质细胞吞噬功能

一种新的体外活体成像法测定星形胶质细胞的吞噬作用 pH 指标-共轭突

巨噬细胞吞噬胸腺细胞吞噬的实验分析

Flow Cytometry Approach to Characterize Phagocytic Properties of Acutely-Isolated Adult Microglia and Brain Macrophages In Vitro

iPSC-巨噬细胞死亡神经母细胞的体外定量成像检测

人小胶质细胞样细胞:使用人突触体从诱导多能干细胞和 体外 活细胞吞噬测定中分化

使用流式细胞术定量突触材料的小胶质细胞吞噬

使用流式细胞术定量突触材料的小胶质细胞吞噬

使用流式细胞术定量突触材料的小胶质细胞吞噬

人小胶质细胞样细胞:使用人突触体从诱导多能干细胞和体外活细胞吞噬测定中分化