新生小鼠视网膜神经节细胞的逆行荧光金标记
Retrograde fluorogold labeling of retinal ganglion cells in neonatal mice.
作者信息
Hu Huiling, Liu Ying, Li Kang, Fang Min, Zou Yunyun, Wang Jiantao, Ge Jian
机构信息
Shenzhen Eye Hospital, Shenzhen Eye Hospital Affiliated to Jinan University, School of Optometry, Shenzhen University, Shenzhen, China.
State Key Laboratory of Ophthalmology, Guangdong Provincial Key Laboratory of Ophthalmology and Visual Science, Zhongshan Ophthalmic Center, Sun Yat-sen University, Guangzhou, China.
出版信息
Ann Transl Med. 2021 May;9(10):878. doi: 10.21037/atm-21-2022.
BACKGROUND
The neonatal period, especially postnatal day 10 (P10), is important for mouse retinal ganglion cells (RGCs) development, and an effective labeling technique to track neonatal RGCs is needed. Retrograde fluorogold (FG) labeling is widely used for adult mouse RGCs, but its applicability for the neonatal mouse is still unknown. This study aimed to evaluate the safety and efficiency of retrograde FG labeling in P10 mice.
METHODS
The anatomic location of the superior colliculus (SC) of P10 wild-type C57/BL6J mice was clarified by histological brain section and hematoxylin and eosin (H&E) staining. Three doses of 3% FG were injected into the SC of 30 mice, and 3 days post-surgery, labeling efficiency was quantified by retinal flat-mounts, and labeling safety was evaluated by mice mortality.
RESULTS
Samples of brain tissue from 2-3.5 mm posterior to the bregma, and from 0.5-2.0 mm lateral to the midline showed major SC-related structures. The FG-positive RGC density in the 0.3 µL group was 3,563.9±311.9 cells/mm, significantly more than in the 0.6 µL group (1,718.6±177.1 cells/mm) or 1.0 µL group (2,496.8±342.2 cells/mm). The mortality rate was 10% in both the 0.3 and 0.6 µL groups, but 40% in the 1.0 µL group.
CONCLUSIONS
The appropriate labeling site in P10 mice was confirmed and 0.3 µL FG is an appropriate dose for retrograde labeling of RGCs.
背景
新生期，尤其是出生后第10天（P10），对小鼠视网膜神经节细胞（RGCs）的发育很重要，因此需要一种有效的标记技术来追踪新生RGCs。逆行荧光金（FG）标记广泛用于成年小鼠RGCs，但它对新生小鼠的适用性仍不清楚。本研究旨在评估逆行FG标记在P10小鼠中的安全性和效率。
方法
通过组织学脑切片和苏木精-伊红（H&E）染色明确P10野生型C57/BL6J小鼠上丘（SC）的解剖位置。将三剂3%的FG注入30只小鼠的SC，术后3天，通过视网膜铺片定量标记效率，并通过小鼠死亡率评估标记安全性。
结果
在脑回后2 - 3.5毫米、中线外侧0.5 - 2.0毫米处的脑组织样本显示出主要的与SC相关的结构。0.3微升组的FG阳性RGC密度为3563.9±311.9个细胞/毫米，显著高于0.6微升组（1718.6±177.1个细胞/毫米）或1.0微升组（2496.8±342.2个细胞/毫米）。0.3微升组和0.6微升组的死亡率均为10%，但1.0微升组为40%。
结论
确定了P10小鼠合适的标记部位，0.3微升FG是RGCs逆行标记的合适剂量。
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