Collection of Frozen Rodent Brain Regions for Downstream Analyses
Collection of Frozen Rodent Brain Regions for Downstream Analyses
复制标题
DOI：
10.3791/60474
复制
发表时间：
2020-04-01
影响因子：
1.2
通讯作者：
Mackie, Ken
中科院分区：
文献类型：
--
作者：
Wager-Miller, James;Green, Michelle Murphy;Mackie, Ken
随着我们对神经生物学的了解的进步，分子分析经常在大脑的小区域进行，如内侧前额叶皮质(MPFC)或伏隔核。这项工作的挑战是在解剖正确区域的同时保护要检查的微环境。在这篇白皮书中，我们描述了一种简单、低成本的方法，该方法利用了大多数实验中现成的资源。这种方法通过在整个过程中保持组织的冰冻来保存核酸和蛋白质。使用大脑基质将大脑切成0.5-1.0 mm的切片，并将其排列在冰冻的玻璃板上。每个部分中的地标都与参考文献(如Allen Mouse Brain Atlas)进行比较，并使用冷刀或活组织检查冲床解剖区域。然后将组织储存在零下80摄氏度直到使用。通过这个过程，大鼠和小鼠的mPFC、伏隔核、背侧和腹侧海马区等区域已经成功地用qRT-PCR和Western分析进行了分析。这种方法仅限于可以通过清晰的地标识别的大脑区域。
As our understanding of neurobiology has progressed, molecular analyses are often performed on small brain areas such as the medial prefrontal cortex (mPFC) or nucleus accumbens. The challenge in this work is to dissect the correct area while preserving the microenvironment to be examined. In this paper, we describe a simple, low-cost method using resources readily available in most labs. This method preserves nucleic acid and proteins by keeping the tissue frozen throughout the process. Brains are cut into 0.5-1.0 mm sections using a brain matrix and arranged on a frozen glass plate. Landmarks within each section are compared to a reference, such as the Allen Mouse Brain Atlas, and regions are dissected using a cold scalpel or biopsy punch. Tissue is then stored at -80 degrees C until use. Through this process rat and mouse mPFC, nucleus accumbens, dorsal and ventral hippocampus and other regions have been successfully analyzed using qRT-PCR and Western assays. This method is limited to brain regions that can be identified by clear landmarks.