肿瘤血管CD34免疫荧光染色条件的探讨与优化  

Exploration and Optimization of CD34 Immunofluorescence Staining Conditions for Tumor Blood Vessels

作　　者：苏孙益 艾栩如 蒋凤琨 廖佳伟 王虹 王一铮[1] 林凡[1] SU Sunyi;AI Xuru;JIANG Fengkun(College of Integrative Medicine,Fujian University of Traditional Chinese Medicine,Fuzhou City,Fujian Province 350122)

机构地区：[1]福建中医药大学中西医结合学院,福建省福州市350122 [2]福建省立医院健康管理科

出　　处：《医学理论与实践》2025年第3期361-365,共5页The Journal of Medical Theory and Practice

基　　金：国家自然科学基金(82074191,82204969);国家大学生创新创业训练计划项目(202310393011);福建省大学生创新创业训练计划项目(S202310393004)。

摘　　要：目的:探讨不同染色条件对肿瘤血管CD34免疫荧光染色结果的影响,为相关研究提供借鉴。方法:选取小鼠Lewis肺癌移植瘤组织石蜡切片,在常规免疫荧光染色基础上,分别使用微波炉、高压锅和水浴锅三种方法修复3min、6min、9min,分别设置3%BSA封闭30min、1h及不封闭组,针对过氧化物酶灭活分别设置抗原修复后封闭前灭活组、一二抗间灭活组和不灭活组,设置稀释比分别为1∶250、1∶500、1∶1000的一抗浓度组及二抗浓度组,分析上述实验条件对肿瘤血管CD34免疫荧光染色结果的影响。结果:抗原修复方法:微波炉组和高压锅组随着修复时间延长,CD34相对亮度逐渐升高(P<0.05或P<0.01);水浴锅组在不同修复时间条件下,CD34相对亮度无显著差异。修复时间为3min、6min时,微波炉组CD34染色亮度均显著高于高压锅组和水浴锅组(P<0.05或P<0.01),修复时间为9min时,微波炉组CD34染色亮度高于水浴锅组(P<0.01),但与高压锅组无显著差异;三个修复时间微波炉组背景亮度均显著高于水浴锅组和高压锅组(P<0.01);修复时间为6min、9min时,高压锅组CD34染色亮度高于水浴锅组(P<0.01),且高压锅组背景亮度与水浴锅组无显著差异。封闭时间:封闭30min组背景亮度显著高于封闭1h组(P<0.05),但显著低于不封闭组(P<0.05),且封闭30min组CD34阳性相对亮度显著高于其他两组(P<0.01)。灭活时序:一二抗间灭活组背景亮度显著低于抗原修复后封闭前灭活组和不灭活组(P<0.01)。抗体浓度:不同浓度一抗孵育组织的CD34阳性相对亮度无显著差异,但CD34一抗浓度稀释比为1∶250时背景亮度明显高于稀释比为1∶500和1∶1000的浓度组(P<0.05);二抗稀释比为1∶250的浓度组CD34阳性相对亮度显著高于稀释比为1∶500和1∶1000浓度组(P<0.05),各二抗浓度组背景相对亮度无显著差异。结论:肿瘤血管CD34免疫荧光染色过程中,延长修复时间可提高微波炉�Objective:To explore the effect of different staining conditions on the immunofluorescence staining results of tumor vascular CD34,and provide reference for related research.Methods:Paraffin sections of Lewis lung cancer transplant tumor tissue from mice were selected.Based on conventional immunofluorescence staining,microwave oven,pressure cooker,and water bath were used to repair for 3 minutes,6 minutes,and 9 minutes,respectively.Three groups were set up:3%BSA blocking for 30 minutes,1 hour,and non blocking groups,respectively.For peroxidase inactivation,groups were set up for antigen repair before blocking,primary and secondary antibody inactivation,and non inactivation groups.The dilution ratios of 1∶250,1∶500,and 1∶1000 for primary and secondary antibody concentration groups were set up,respectively.The effects of the above experimental conditions on the immunofluorescence staining results of tumor vascular CD34 were analyzed.Results:Antigen repair methods:With the prolongation of repair time,the relative brightness of CD34 gradually increased in the microwave oven group and pressure cooker group(P<0.05 or P<0.01).There was no significant difference in the relative brightness of CD34 in the water bath group under different repair time conditions.When the repair time was 3 minutes and 6 minutes,the brightness of CD34 staining in the microwave group was significantly higher than that in the pressure cooker group and water bath group(P<0.05 or P<0.01).When the repair time was 9 minutes,the brightness of CD34 staining in the microwave group was higher than that in the water bath group(P<0.01),but there was no significant difference compared to the pressure cooker group.The background brightness of the microwave group was significantly higher than that of the water bath group and pressure cooker group at all three repair times(P<0.01).When the repair time was 6 minutes and 9 minutes,the brightness of CD34 staining in the pressure cooker group was higher than that in the water bath group(P<0.01),and there wa

关 键 词：免疫荧光 条件优化 肿瘤血管 CD34 

分 类 号：Q95-33[生物学—动物学]