四种不同固定液固定小鼠视网膜效果的比较研究

Comparative study of mouse retinal fixation outcomes with four different fixation solutions

目的研究四种不同固定液固定小鼠视网膜石蜡切片的效果。方法将取自6只正常C57/BL6J小鼠的12只眼球,随机分成4组(n=3),置于四种不同的眼球固定液中固定过夜,A组为改良固定液,B组为40 g·L^(- 1)多聚甲醛固定液,C组为Kolmer固定液,D组为Boun固定液,然后按照常规石蜡切片步骤进行标本制作,行HE染色,比较固定和染色效果,同时进行免疫荧光组织化学的比较研究。结果大体观察A组、D组眼球形态饱满不变形,B组、C组眼球有皱缩变形现象。HE染色示,A组标本视网膜细胞排列紧密,各层结构清晰完整,色泽艳丽;B组、C组和D组标本视网膜均有变形、分离等现象。免疫荧光染色示,A组标本视网膜水平细胞和双极细胞显示效果良好,但是无法有效标记出视网膜神经节细胞,而B组标本视网膜可以清晰标记出视网膜神经节细胞。结论改良眼球固定液固定效果优于其他三种固定液,对水平细胞和双极细胞的相关研究较适用,而对于视网膜神经节细胞的相关研究不适合。

Objective To investigate the fixation outcomes of mouse retina paraffin sections with four different fixation solutions. Methods Eyeballs were harvested from six C 57 BL /6J mice and randomly divided into four groups,3 cases in each group,group A（ retina fixation with improved fixation solution）,group B（ retina fixation with 40 g·L（- 1）paraformaldehyde）,group C（ retina fixation with Kolmer solution）,and group D（ retina fixation with Boun solution,n =3）. Paraffin sections were prepared 24 hours after retina fixation,HE staining and immunofluorescence technique were performed,and the outcomes of fixation and staining were compared. Results In general,the shapes of eyeballs were well and nondeformable in group A and D,but were shrunk and deformable in group B and C. HE staining show ed that the retinal structures of group A were clear and intact,but retinal detachment happened in group B and C. Immunofluorescent staining show ed that bipolar cells and horizontal cells could be marked clearly with their specific markers in group A,but retinal ganglion cells could not be labeled effectively. In group B,retinal ganglion cells could be labeled with their specific marker. Conclusion Eyeball fixation with improved fixation solution may result in better outcomes than other three fixation solutions,and is suitable for further research with retinal paraffin sections,better for bipolar cells and horizontal cells,but not suitable for retinal ganglion cells.