Improved method for combination of immunocytochemistry and Nissl staining.
Improved method for combination of immunocytochemistry and Nissl staining.
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DOI：
10.1016/j.jneumeth.2009.07.010
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发表时间：
2009-10-30
影响因子：
3
通讯作者：
Fekete, Csaba
中科院分区：
文献类型：
--
作者：
Kadar, Andrea;Wittmann, Gabor;Liposits, Zsolt;Fekete, Csaba
nisl染色是一种广泛应用于神经组织形态学和病理学研究的方法。经标准免疫细胞化学染色后，nisl染色仅标记神经元核，而神经元核周的特征性染色缺失或很弱。我们假设免疫细胞化学处理过程中的RNA降解导致了nissl染色细胞质的丢失。为了验证这一假设，我们在免疫染色的所有步骤中都使用了无rna酶的条件。为了进一步防止rna被RNAse污染降解，在所有含抗体的溶液中加入RNAse抑制剂肝素。用c-Fos抗体和神经肽Y （NPY）抗体对禁食3 d的大鼠组织进行标准免疫细胞化学和无rnase双标记免疫细胞化学后的nsll染色效率进行比较。经过标准的免疫细胞化学，nisl染色标记了神经元的细胞核，只有非常微弱的细胞质。无rnase处理未改变免疫反应信号的分布，但保留了nissl -染料对神经元核周的染色。综上所述，在免疫细胞化学过程中无rnase的条件下，可以用nissl染料标记神经元核周。所描述的方法有助于免疫细胞化学信号的定位，并使通过其核蛋白含量鉴定神经元的神经支配的光镜检查成为可能。
Nissl-staining is a widely used method to study morphology and pathology of neural tissue. After standard immunocytochemistry, the Nissl-staining labels only the nucleus of neurons and the characteristic staining of the neuronal perikarya is absent or very weak. We hypothesized that the RNA degradation during the immunocytochemical treatment results in the loss of cytoplasmic staining with Nissl-dyes. To test this hypothesis, we used RNAse-free conditions for all steps of immunostaining. To further prevent the RNA-degradation by RNAse contaminations, the RNAse inhibitor heparin was added to all antibody-containing solutions. The efficiency of Nissl-staining after standard and RNAse-free double-labeling immunocytochemistry was compared using antibodies against c-Fos and neuropeptide Y (NPY) on tissues of rats refed after three days of fasting. After standard immunocytochemistry, the Nissl-staining labeled the nuclei of neurons and only very faintly the cytoplasm of these cells. The RNAse-free treatment did not alter the distribution of immunoreaction signal, but preserved the staining of neuronal perikarya by the Nissl-dyes. In conclusion, the RNAse-free conditions during immunocytochemistry, allows the labeling of neuronal perikarya by Nissl-dyes. The described method facilitates the mapping of immunocytochemical signals and makes possible the light microscopic examination of the innervation of neurons identified by their nuclear protein content.
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影响因子：
3.2
作者：
Pilati, Nadia;Barker, Matthew;Hamann, Martine
通讯作者：
Hamann, Martine
影响因子：
2.8
作者：
Kulesza, Randy J., Jr.
通讯作者：
Kulesza, Randy J., Jr.
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影响因子：
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作者：
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通讯作者：
Mushinski JF
影响因子：
12.7
作者：
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通讯作者：
Kusaka, Hirofumi
影响因子：
2.5
作者：
Lorenzo, Louis-Etienne;Ramien, Michele;Ribeiro-Da-Silva, Alfredo
通讯作者：
Ribeiro-Da-Silva, Alfredo