Boone Deborah R, Weisz Harris A, Sell Stacy L, Hellmich Helen L
Department of Anesthesiology, University of Texas Medical Branch, Galveston, TX, USA.
Methods Mol Biol. 2018;1723:235-245. doi: 10.1007/978-1-4939-7558-7_13.
The methods presented here are based on our laboratory's 15 years of experience using laser capture microdissection to obtain samples for the study of gene expression after traumatic brain injury (TBI) using a well-established rat model of experimental TBI. Here, we describe how to use the Arcturus laser capture microdissection system to capture swaths of specific regions of the rat hippocampus as well as specific neuronal populations defined by Fluoro-Jade C staining. Staining with Fluoro-Jade C identifies a neuron that is in the process of degeneration. We have optimized our protocols for Fluoro-Jade C tissue staining and laser capture microdissection to maintain RNA integrity which is essential for a variety of downstream applications, such as microarray, PCR array, and quantitative real-time PCR analyses.
本文介绍的方法基于我们实验室15年的经验，即使用激光捕获显微切割技术，通过一个成熟的实验性创伤性脑损伤（TBI）大鼠模型获取样本，以研究创伤性脑损伤后的基因表达。在这里，我们描述了如何使用Arcturus激光捕获显微切割系统来捕获大鼠海马特定区域的条带，以及由Fluoro-Jade C染色定义的特定神经元群体。用Fluoro-Jade C染色可识别正在发生变性的神经元。我们已经优化了Fluoro-Jade C组织染色和激光捕获显微切割的方案，以保持RNA完整性，这对于各种下游应用（如微阵列、PCR阵列和定量实时PCR分析）至关重要。
