石蜡切片多重免疫荧光染色优化条件探讨  

Discussion of Multiple Immunofluorescence Staining Optimized Conditions on Paraffin Section

作　　者：范瑾瑾[1] 骆宁[1] 彭文兴[1] 毛海萍[1] FAN Jin-jin;LUO Ning;PENG Wen-xing;MAO Hai-ping(Department of Nephrology,The First Affiliated Hospital,Sun Yat-sen University,Guangzhou 510080,China)

机构地区：[1]中山大学附属第一医院肾内科,广东广州510080

出　　处：《中山大学学报（医学版）》2018年第5期675-681,共7页Journal of Sun Yat-Sen University:Medical Sciences

基　　金：广东省肾脏病重点实验室基金(214B030301023);广东省医学科学基金(A2018042)

摘　　要：【目的】探讨在石蜡切片上不同染色条件对多重荧光标记结果的影响。【方法】根据不同组织抗原选择合适的固定剂;对石蜡组织切片采用高温高压的抗原修复方式,使用柠檬酸盐缓冲液或EDTA抗原修复液进行抗原修复,并在染色操作过程中增加封闭步骤,随后进行常规组织免疫荧光染色。【结果】常规石蜡切片用40g/L多聚甲醛固定、抗原糖基含量高的石蜡切片用过碘酸盐-赖氨酸-多聚甲醛固定可以很好的保护切片中的抗原;pH 6.0的枸橼酸缓冲液及pH 8.0的EDTA-Na缓冲液分别对组织胞浆抗原及胞核抗原具有较好的抗原修复作用;多重荧光染色不同种属来源第一抗体染色间增加一次封闭步骤,可有效的改善多重荧光染色质量。【结论】选择合适的固定剂、抗原修复液及增加抗原封闭次数对石蜡组织切片上的多重荧光标记结果具有显著的影响。【Objective】To investigate the effect of different multiple immunostaining condition on tissue paraffin section.【Methods】Tissue paraffin sections were treated with different fixing agents according to thediverse tissue antigens.The tissue antigens were retrieved with citrate buffer or EDTA-Na buffer by using high temperature and high pressure antigen retriever.Additional blocking processing was performed following by the conventional tissue immunofluorescence staining.【Results】The citrate buffer(pH 6.0)was suited for cytoplasm antigens retrieve,nevertheless the EDTA-Na buffer(pH 8.0)was fitted fornuclear antigens.For the multiple immunofluorescence staining,additional block processing between the various species first antibodies staining can significant improve the staining results.【Conclusions】Suitable fixing agent,matched antigen retrieve buffer and additional blocking process were critical for getting perfect results in multiple immunofluorescence staining on tissue paraffin section.

关 键 词：多重免疫荧光染色 石蜡切片 优化条件 

分 类 号：R331[医药卫生—人体生理学]