Triple fluorescence labelling of neuronal, glial and vascular markers revealing pathological alterations in various animal models
Triple fluorescence labelling of neuronal, glial and vascular markers revealing pathological alterations in various animal models
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DOI：
10.1016/j.jchemneu.2008.10.003
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发表时间：
2009-03-01
影响因子：
2.8
通讯作者：
Grosche, Jens
中科院分区：
文献类型：
--
作者：
Haertig, Wolfgang;Reichenbach, Andreas;Grosche, Jens
同时检测神经胶质细胞、血管和神经元有助于深入了解中枢神经系统复杂的化学结构。在此，我们提出一种简单、可靠且通用的方法，用于对神经元、血管和神经胶质标记物进行羰花青三重荧光标记。该方法在生理条件下的大鼠脑组织、控制性皮质撞击损伤导致的创伤性脑损伤后以及大脑中动脉闭塞后的中风模型中显示出其有效性。此外，通过将其应用于具有年龄相关性β -淀粉样变和海马体中tau蛋白过度磷酸化的老年三重转基因小鼠的切片，验证了该方法的通用性，这些小鼠模拟了阿尔茨海默病的神经病理改变。为了举例说明该方法在分析肠神经系统方面的有效性，将其应用于马肠道的整装片。来自马铃薯（Solanum tuberosum）的生物素化凝集素是一种检测血管和小胶质细胞的极佳工具。此外，这种凝集素在实验性损伤后可显示巨噬细胞，在老年三重转基因小鼠中可显示老年斑。通过对胶质纤维酸性蛋白进行免疫标记证明了大部分星形胶质细胞的存在。通过针对神经元核的单克隆抗体检测神经元，在马组织中，使用识别一种保守核蛋白的小鼠抗 - HuC/D抗体进行检测。共聚焦激光扫描显微镜阐明了相关标记物的空间关系以及在实验性损伤后和具有阿尔茨海默样病变的转基因小鼠中的病理改变。（C）2008爱思唯尔有限公司。保留所有权利。
The simultaneous detection of glia, vessels and neurons facilitates insights into the complex chemoarchitecture of the central nervous system. Here, we present a simple, robust and versatile approach for the carbocyanine triple fluorescence labelling of neuronal, vascular and glial markers.The usefulness of this procedure is shown for rat brain tissue under physiological conditions, after traumatic brain injury caused by controlled cortical impact injury, and after stroke following middle cerebral artery occlusion. Moreover, the versatility of the method is verified by its application to sections from old triple transgenic mice with age-dependent P-amyloidosis and tau hyperphosphorylation in the hippocampus, modelling neuropathological alterations in Alzheimer's disease. To exemplify the usefulness of the approach for analysis of the enteric nervous system, it was applied to whole mounts from the horse intestine.The biotinylated lectin from potato (Solanum tuberosum) is presented as an excellent tool to detect both vessels and microglia. Furthermore, this lectin revealed macrophages after experimental insults, and senile plaques in aged triple transgenic mice. A large portion of astroglia was demonstrated by immunolabelling of glial fibrillary acidic protein. Neurons were detected by monoclonal antibodies directed against neuronal nuclei and, in horse tissues, mouse-anti-HuC/D recognizing a conserved nuclear protein. Confocal laser-scanning microscopy elucidated spatial relationships of the relevant markers and their pathological alterations after experimental insults and in transgenic mice with Alzheimer-like lesions. (C) 2008 Elsevier B.V. All rights reserved.