小鼠胰腺石蜡切片制作程序优化与免疫荧光染色探索  

Optimization of the procedure for paraffin sections preparation procedure of the mouse pancreas and exploration of immunofluorescence staining

作　　者：赵晶晶 庄宝祥[2] 李如江[1] Zhao Jingjing;Zhuang Baoxiang;Li Rujiang(Department of Histology and Embryology,Weifang Medical University,Weifang 261053,China;Department of Morphological Laboratory,Weifang Medical University,Weifang 261053,China)

机构地区：[1]潍坊医学院组织学与胚胎学教研室,山东省高校重点实验室,潍坊261053 [2]潍坊医学院组织学与胚胎学教研室潍坊医学院形态学实验室,潍坊261053

出　　处：《中国组织化学与细胞化学杂志》2019年第4期384-387,共4页Chinese Journal of Histochemistry and Cytochemistry

基　　金：国家自然科学基金(81471048);山东省高等学校科技计划项目(J12LK05)

摘　　要：目的优化小鼠胰腺石蜡切片的制作程序并分析1型糖尿病小鼠胰腺免疫荧光染色结果。方法多次小剂量链脲佐菌素诱导小鼠1型糖尿病,取胰腺,观察采用不同试剂和处理时间的石蜡包埋程序对胰腺切片质量的影响,并探讨胰腺石蜡切片免疫荧光染色的可能假阳性。结果95%乙醇15min×2次、100%乙醇Ⅰ6min、100%乙醇Ⅱ7min、苯Ⅰ15min、苯Ⅱ18min、浸蜡90min时制作的小鼠胰腺石蜡切片质量最优。以小鼠抗高血糖素抗体(一抗)及FITC-山羊抗小鼠二抗免疫染色1型糖尿病小鼠胰腺时常常出现假阳性荧光。结论脱水、透明、浸蜡等制作程序对小鼠胰腺石蜡切片质量影响较大;苯是优于氯仿与二甲苯的胰腺透明剂;1型糖尿病小鼠胰腺不适合以小鼠源抗体为一抗进行间接免疫荧光染色。Objective To optimize the paraf fin sections preparation procedure of mouse pancreas and analyze the results of immunofluorescence staining of pancreas in type 1 diabetic mice.Methods Repeated low doses of streptozotocin were used to induce mice model with type 1 diabetes mellitus.The pancreas were taken to explore the effects of different time and reagents for dehydration,transparency,paraf fin immersion and paraf fin embedding on the quality of pancreatic paraf fin sections,and to explore the mechanism of possible immuno fluorescent false positive staining in paraf fin sections of pancreas.Results The best quality of mice pancreatic par-af fin sections was obtained by adopting 95%ethanol 15 min×2 times,100%ethanol I 6 min,100%ethanol II 7 min,benzene I 15 min,benzene II 18 min and wax immersion 90 min.False-positive fluorescence staining was observed when the pancreas of type 1 diabetic mice were stained with mouse anti-Glucagon primary antibody and FITC-conjugated goat-anti-mouse secondary antibody.Conclusion Procedures such as dehydration,transparency and wax immersion have a great impact on the quality of paraf fin sections of pancreas from mice.Benzene is superior to chloroform and xylene as pancreatic transparent agent.Mouse-derived antibody are not suitable as primary antibody for indirect immunofluorescent staining in pancreas of type 1 diabetic mice.

关 键 词：石蜡切片 胰腺 免疫荧光染色 非特异性染色 

分 类 号：R361[医药卫生—病理学]