Children's Hospital Los Angeles, Los Angeles, CA 90027, USA.
University of Southern California, Los Angeles, CA 90007, USA.
Cell Rep Methods. 2022 Oct 14;2(10):100316. doi: 10.1016/j.crmeth.2022.100316. eCollection 2022 Oct 24.
Spatial gene expression, achieved classically through hybridization, is a fundamental tool for topographic phenotyping of cell types in the nervous system. Newly developed techniques allow for visualization of multiple mRNAs at single-cell resolution and greatly expand the ability to link gene expression to tissue topography, yet there are challenges in efficient quantification and analysis of these high-dimensional datasets. We have therefore developed the single-cell automated multiplex pipeline for RNA (SCAMPR), facilitating rapid and accurate segmentation of neuronal cell bodies using a dual immunohistochemistry-RNAscope protocol and quantification of low- and high-abundance mRNA signals using open-source image processing and automated segmentation tools. Proof of principle using SCAMPR focused on spatial mapping of gene expression by peripheral (vagal nodose) and central (visual cortex) neurons. The analytical effectiveness of SCAMPR is demonstrated by identifying the impact of early life stress on gene expression in vagal neuron subtypes.
空间基因表达，通过杂交经典实现，是对神经系统中细胞类型进行地形表型分析的基本工具。新开发的技术允许在单细胞分辨率下可视化多个 mRNA，并极大地扩展了将基因表达与组织地形联系起来的能力，但在高效量化和分析这些高维数据集方面存在挑战。因此，我们开发了单细胞自动化多重 RNA 分析 (SCAMPR) 方法，该方法使用双重免疫组织化学-RNAscope 方案促进神经元细胞体的快速和准确分割，并使用开源图像处理和自动分割工具对低丰度和高丰度 mRNA 信号进行定量。使用 SCAMPR 进行原理验证的重点是通过外周（迷走神经结）和中枢（视觉皮层）神经元进行基因表达的空间映射。SCAMPR 的分析效果通过识别早期生活应激对迷走神经元亚型基因表达的影响得到证明。
