Immunoblotting patterns of cytoskeletal dendritic protein expression in human neocortex
Immunoblotting patterns of cytoskeletal dendritic protein expression in human neocortex
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DOI：
10.1007/bf02815127
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发表时间：
1997-08-01
期刊：
影响因子：
--
通讯作者：
Lishaa, NA
中科院分区：
文献类型：
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作者：
Kaufmann, WE;Taylor, CV;Lishaa, NA
细胞骨架蛋白的定性和定量评估对于了解影响神经系统的生理和病理过程至关重要。大多数针对人类样本的此类研究仅使用免疫组织化学技术。我们描述了一种用于评估神经元细胞骨架蛋白的互补免疫印迹方法，该方法采用新鲜冷冻的死后组织。我们发现胞质组分适合对四种主要树突状细胞骨架蛋白进行定性和定量评估：微管相关蛋白 (MAP)-2、MAP-5 和高分子量和中等分子量非磷酸化神经丝。增强化学发光 (ECL) 技术揭示了猴子（无死后延迟）和人类（死后间隔 17-34 小时）样本中所有四种蛋白质的一致且独特的免疫印迹模式，其中一些与啮齿类动物中发现的不同。通过组织蛋白-光密度曲线进行的印迹定量显示了 0 至 100 μg 范围内测量的线性，支持这些免疫测定法用于研究神经系统疾病的可行性。
Qualitative and quantitative evaluations of cytoskeletal proteins are critical for understanding physiological and pathological processes affecting the nervous system. Most of such studies on human samples have only used immunohistochemical techniques. We describe a complementary immunoblotting approach, for the assessment of neuronal cytoskeletal proteins, which employs fresh frozen postmortem tissues. We found that cytosolic fractions are suitable for qualitative and quantitative evaluations of four major dendritic cytoskeletal proteins: microtubule-associated protein (MAP)-2, MAP-5, and high- and medium-molecular-weight nonphosphorylated neurofilaments. The enhanced chemiluminescence (ECL) technique revealed consistent and distinctive immunoblotting patterns for all four proteins in both monkey (no postmortem delay) and human (17-34 h postmortem interval) samples, some of which differed from those found in rodents. Quantitations of blots, by tissue protein-optical density curves that demonstrated linearity of the measurements in the 0- to 100-mu g range, support the feasibility of these immunoassays for the study of neurologic disorders.