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Author:
Date:
2014-02-05
[Abstract] We describe a water-based optical clearing agent, SeeDB (See Deep Brain), which clears fixed brain samples in a few days without quenching many types of fluorescent dyes, including fluorescent proteins and lipophilic neuronal tracers. SeeDB is a saturated solution of fructose (80.2% w/w) in water with 0.5% α-thioglycerol. In standard SeeDB optical clearing procedure, we treat paraformaldehyde-fixed embryo and brain samples with increasing concentrations of aqueous fructose solutions, and finally equilibrate them in SeeDB. The entire procedure takes approximately three days. Unlike previous methods, this method maintains a constant sample volume during the clearing procedure, an important factor to keep cellular morphology intact. After optical clearing, we can reach > 1,000 μm under ...
[摘要] 我们描述了一种水基光学清除剂，SeeDB（参见深脑），其在几天内清除固定的脑样品，而不淬灭许多类型的荧光染料，包括荧光蛋白和亲脂性神经元示踪剂。 SeeDB是果糖（80.2％w/w）在含有0.5％α-硫代甘油的水中的饱和溶液。在标准的SeeDB光学清除程序中，我们用增加浓度的果糖水溶液处理多聚甲醛固定的胚胎和脑样品，并且最后在SeeDB中平衡它们。整个过程大约需要三天。与以前的方法不同，该方法在清除过程中保持恒定的样品体积，这是保持细胞形态完整的重要因素。光学清除后，我们可以达到>在共聚焦显微镜下为1,000μm。当与双光子显微镜结合时，SeeDB允许我们以毫米级别水平成像固定的小鼠大脑。这种方法有助于全面和定量分析理解神经元电路，在成人和发展中国家的小鼠大脑。还提供了SeeDB变体（参见DB37）和优化的程序（见DBp和SeeDB37ht协议）以满足特定要求。
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