Department of Biology, University of Nevada, Reno, Nevada 89557, USA
Department of Biological Sciences, University of Arkansas, Fayetteville, Arkansas 72701, USA
Cold Spring Harb Protoc. 2024 Sep 3;2024(9):pdb.prot108117. doi: 10.1101/pdb.prot108117.
The technique of visualizing axon pathways in the embryonic ventral nerve cord using antibody labeling has been fundamental to our understanding of the genetic and developmental mechanisms underlying nervous system wiring in High-resolution microscopic examination of the ventral nerve cord remains an essential component of many experiments in developmental neuroscience. Although it is possible to examine the ventral nerve cord in intact whole-mount embryos, to collect the highest-quality images it is often useful to isolate the nervous system away from the other embryonic tissues through embryo dissection. This protocol describes methods for dissecting ventral nerve cords from embryos that have been fixed and stained via immunofluorescence or horseradish peroxidase (HRP) immunohistochemistry. The process of making fine dissection needles for this purpose from electrolytically sharpened tungsten wire is also described. Dissected and mounted ventral nerve cords can be examined and imaged using a variety of microscopy techniques including differential interference contrast (DIC) optics, epifluorescence, or confocal microscopy.
使用抗体标记可视化胚胎腹神经索轴突途径的技术是我们理解神经系统布线的遗传和发育机制的基础。高分辨率显微镜检查腹神经索仍然是许多发育神经科学实验的基本组成部分。虽然可以在完整的全胚胎中检查腹神经索，但为了收集最高质量的图像，通常需要通过胚胎解剖将神经系统与其他胚胎组织分离。本协议描述了从已通过免疫荧光或辣根过氧化物酶 (HRP) 免疫组织化学固定和染色的胚胎中分离腹神经索的方法。还描述了为此目的从电解锐化钨丝制作精细解剖针的过程。可以使用各种显微镜技术检查和成像解剖和安装的腹神经索，包括微分干涉对比 (DIC) 光学、荧光和共聚焦显微镜。
