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首页> 外文期刊>Journal of Neuroscience Methods >A quantitative method for analysis of in vitro neurite outgrowth.
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A quantitative method for analysis of in vitro neurite outgrowth.

机译:体外神经突生长分析的定量方法。

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The adult mammalian CNS is extremely limited in its ability to regenerate axons following injury. Glial scar, neuroinflammatory processes and molecules released from myelin impair axonal regrowth and contribute to the lack of neural regeneration. An in vitro assay that quantitates neurite outgrowth from cultured neurons as a model of neuronal regenerative potential is described. Specifically, the neurite outgrowth from primary neurons (rat cerebellar granule neurons; CGNs) and a neuronal cell line (NG108-15) were quantitatively measured after optimization of culture conditions. After cultures were fixed and immunostained to label neurons and nuclei, microscope images were captured and an image analysis algorithm was developed using Image-Pro Plus software to allow quantitative analysis. The algorithm allowed the determination of total neurite length, number of neurons, and number of neurons without neurites. The algorithm also allows for end-user control of thresholds for staining intensity and celluclei size. This assay represents a useful tool for quantification of neurite outgrowth from a variety of neuronal sources with applications that include: (1) assessment of neurite outgrowth potential; (2) identification of molecules that can block or stimulate neurite outgrowth in conventional culture media; and (3) identification of agents that can overcome neurite outgrowth inhibition by inhibitory substrates.
机译:成年哺乳动物中枢神经系统在损伤后其再生轴突的能力极其有限。胶质瘢痕,神经炎性过程和髓磷脂释放的分子会损害轴突再生,并导致缺乏神经再生。描述了一种体外测定,该测定定量了作为神经元再生潜能模型的来自培养的神经元的神经突生长。具体而言,在优化培养条件后,定量测量了初级神经元(大鼠小脑颗粒神经元; CGN)和神经元细胞系(NG108-15)的神经突生长。固定培养物并免疫染色以标记神经元和细胞核后,捕获显微镜图像,并使用Image-Pro Plus软件开发图像分析算法以进行定量分析。该算法可以确定神经突的总长度,神经元数量以及没有神经突的神经元数量。该算法还允许最终用户控制染色强度和细胞/细胞核大小的阈值。该测定法是从多种神经元来源定量神经突增生的有用工具,其应用包括:(1)评估神经突增生潜力; (2)鉴定可阻止或刺激常规培养基中神经突生长的分子; (3)鉴定能克服抑制底物抑制神经突生长的药物。

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