Quantifying synapses: an immunocytochemistry-based assay to quantify synapse number.
Quantifying synapses: an immunocytochemistry-based assay to quantify synapse number.
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DOI：
10.3791/2270
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发表时间：
2010-11-16
期刊：
影响因子：
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通讯作者：
Eroglu, Cagla
中科院分区：
文献类型：
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作者：
Ippolito, Dominic M;Eroglu, Cagla
神经科学最重要的目标之一是了解指导突触形成早期阶段的分子线索。因此，必须开发客观的方法来量化突触连接的变化。从样品固定开始，该方案详细介绍了如何使用免疫细胞化学定量分离的神经元培养物和脑切片中的突触数量。使用隔室特异性抗体，我们标记突触前终端以及网站的突触后专业化。我们将突触定义为这些标记物产生的信号之间的共定位点。这些共定位的数量使用ImageJ分析软件平台下的Puncta Analyzer（由Bary Wark编写，可根据要求获得，c.eroglu@www.example.com）中的插件进行定量。cellbio.duke.edu本方案中描述的突触测定可应用于任何神经组织或培养制备物，您可以选择突触前和突触后标记物。这种突触测定是一种有价值的工具，可广泛用于突触发育的研究。
One of the most important goals in neuroscience is to understand the molecular cues that instruct early stages of synapse formation. As such it has become imperative to develop objective approaches to quantify changes in synaptic connectivity. Starting from sample fixation, this protocol details how to quantify synapse number both in dissociated neuronal culture and in brain sections using immunocytochemistry. Using compartment-specific antibodies, we label presynaptic terminals as well as sites of postsynaptic specialization. We define synapses as points of colocalization between the signals generated by these markers. The number of these colocalizations is quantified using a plug in Puncta Analyzer (written by Bary Wark, available upon request, c.eroglu@cellbio.duke.edu) under the ImageJ analysis software platform. The synapse assay described in this protocol can be applied to any neural tissue or culture preparation for which you have selective pre- and postsynaptic markers. This synapse assay is a valuable tool that can be widely utilized in the study of synaptic development.