不同固定液对豚鼠免疫器官石蜡切片质量的影响

Effects of Different Fixative Fluids on the Quality of Paraffin Sections in Immune Organs of Guinea Pigs

目的:比较不同固定液对SPF级豚鼠免疫器官的固定效果,为筛选适用于脾脏、淋巴结和骨髓的固定液提供参考。方法:将21只300~350 g雄性SPF级豚鼠随机分为7组,每组3只,摘取脾脏、肠系膜淋巴结、髂骨骨髓,各组分别使用4%多聚甲醛溶液、10%中性福尔马林溶液、Bouin’s溶液、Carnoy溶液、Davidson’s溶液、Zenker溶液、Helly溶液固定48 h。髂骨骨髓在固定前浸泡于EDTA脱钙液(pH7.2)中并置于37℃孵箱进行脱钙。通过制作石蜡切片并进行HE染色,从切片龟裂程度、细胞形态、染色效果等方面比较7种固定液对切片质量的影响。结果:10%中性福尔马林溶液固定后的脾脏未出现龟裂,白髓和红髓着色清晰,淋巴细胞形态易辨。Carnoy溶液固定后的淋巴结生发中心明区暗区分明,红蓝染色适中,淋巴细胞形态清晰。4%多聚甲醛溶液固定后的骨髓着色适中,清晰可辨。结论:常温条件下对组织固定48 h,经HE染色后想要达到理想的切片质量,建议使用10%中性福尔马林溶液固定脾脏,使用Carnoy溶液固定淋巴结,首选4%多聚甲醛溶液固定骨髓,如果考虑固定的同时完成脱钙建议选择Bouin’s溶液固定骨髓。

Objective:To investigate the fixation effect of different fixatives on immune organs of SPF grade guinea pigs,and provide reference for screening fixatives suitable for spleen,lymph nodes and bone marrow.Methods:Twenty-one 300-350 g SPF male guinea pigs were randomly divided into 7 groups with 3 in each group.Spleen,mesenteric lymph nodes and iliac bone marrow were extracted.Each group was fixed with 4%polyformaldehyde solution,10%neutral formalin solution,Bouin's solution,Carnoy solution,Davidson's solution,Zenker solution and Helly solution for 48 h,respectively.Before fixation,iliac bone marrow was immersed in EDTA decalcification solution(pH7.2)and placed in a incubator at 37℃for decalcification.By making paraffin sections and staining with HE,the effects of 7 fixative fluids on section quality were compared from the aspects of section cracking degree,cell morphology and staining effect.Results:The results showed that the white pulp and red pulp of spleen after 10%neutral formalin solution were clearly colored,no cracks appeared,and the morphology of lymphocytes was easy to distinguish.After fixation with Carnoy solution,the bright and dark areas of the germinal center of the lymph nodes were distinct,the red and blue staining was moderate,and the lymphocyte morphology was clear.After fixation with 4%paraformaldehyde solution,bone marrow staining was moderate and clearly distinguishable.Conclusions:For tissue fixation at room temperature for 48 h,after HE staining,it is recommended to use 10%neutral formalin solution to fix the spleen,Carnoy solution to fix the lymph nodes,and 4%paraformaldehyde solution to fix the bone marrow,and Bouin's solution to fix the bone marrow if decalcification while fixation is considered.