XU Te,,et al.Technique improvement of multiple immunohistochemical staining on tissue microarray[J].Journal of Shenzhen University Science and Engineering,2019,36(5):564-569.[doi:10.3724/SP.J.1249.2019.05564]





Technique improvement of multiple immunohistochemical staining on tissue microarray
4)汕头大学医学院附属肿瘤医院,广东汕头 515000
XU Te1 2 3 WANG Yun1 DENG Fang1 4 LI Yanpeng1 QU Junle2 YAN Hongfei4 and SHENG Sitong1
1) College of Life Sciences and Oceanography, Shenzhen University, Shenzhen 518055, Guangdong Province, P.R.China
2) College of Physics and Optoelectronic Engineering, Key Laboratory of Optoelectronic Devices and Systems of Ministry of Education and Guangdong Province, Shenzhen University, Shenzhen 518060, Guangdong Province, P.R.China
3) College of Stomatology, Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research, Xi’an Jiaotong University, Xi’an 710004, Shaanxi Province, P.R.China
4) Cancer Hospital, Shantou University Medical School, Shantou 515000, Guangdong Province, P.R.China
molecular pathology tissue microarray immunohistochemistry multiple staining antigen retrieval
为提高多重免疫组织化学(immunohistochemistry, IHC)技术在组织芯片(tissue microarray, TMA)上的染色效果,通过对同一样本进行多种抗体染色或同种抗体多次染色的方法,探索并优化IHC技术中的多个步骤,具体包括:“滤纸夹层-蒸蛋器”抗原修复新装置的研发、抗原修复液的筛选、透明剂的优选. 研究结果显示:自创的“滤纸夹层-蒸蛋器”抗原修复辅助工具,能减少高温高压修复时水分子对组织的冲击,作用温和、操作安全,有较好的组织切片完整性和较少的组织脱片率,且可减少抗原修复液使用量;柠康酐抗原修复液对多重染色具有更强的阳性效应和通用性;无毒TO(terpentine oil)透明剂可取代有毒二甲苯,具有同样的脱蜡透明效果.TMA中多重IHC技术的改良,提供了更加高效、廉价、安全的方法,可提升与扩展TMA在病理学中的应用价值.
To improve the technique of multiple immunohistochemistry (IHC) on tissue microarray (TMA), several steps of multiple IHC are optimized via utilizing multi-antibody or single antibody multi-staining in one microarray. The optimized methods include “filter paper interlayer-egg steamer” antigen retrieval tool, antigen retrieval buffer and transparent reagent. The results show that, using invented set of “filter paper interlayer-egg steamer”, it can mitigate the impact force of water flow during antigen retrieval process to protect tissue damage, and save antigen retrieval buffer. As antigen retrieval solution, citraconic anhydride has stronger positive effect on multiple staining and for most antibodies. As transparent solution, non-toxic terpentine oil (TO) has the same results of deparaffin and transparent effects. In conclusion, the improve of multiple IHC provides a more efficient, cheap and safe method, which can greatly enhance the value of TMA application in clinical pathology.


[1] TORLAKOVIC E E, NIELSEN S, VYBERG M, et al. Getting controls under control: the time is now for immunohistochemistry[J]. Journal of Clinical Pathology, 2015, 68(11): 879-882.
[2] SHI Shanrong, SHI Yan, TAYLOR C R. Antigen retrieval in immunohistochemistry: review and future prospects in research and diagnosis over two decades[J]. Journal of Histochemistry & Cytochemistry, 2011, 59(1): 13-32.
[3] BOGEN S A, VANI K, SOMPURAM S R. Molecular mechanisms of antigen retrieval: antigen retrieval reverses steric interference caused by formalin-induced cross-links[J]. Biotechnic & Histochemistry, 2009, 84(5): 207-215.
[4] BOGEN S A, VANI K, SOMPURAM S R. Molecular mechanisms of antigen retrieval: antigen retrieval reverses steric interference caused by formalin-induced cross-links[J]. Biotechnic & Histochemistry: Official Publication of the Biological Stain Commission, 2009, 84(5): 207-215.
[5] ZHAO Cai, JIM M, KATHY C, et al. Immunohistochemical staining of epoxy resin sections of peripheral nerve[J]. Applied Immunohistochemistry & Molecular Morphology, 2005, 13(3): 292-294.
[6] 许特,李延鹏,赵宋礼,等.人结直肠癌中两种新分子生物标志物的筛选[J].深圳大学学报理工版,2018,35(5):529-535.
XU Te, LI Yanpeng, ZHAO Songli, et al. Two new molecular biomarkers in human colorectal cancer[J]. Journal of Shenzhen University Science and Engineering, 2018, 35(5): 529-535.(in Chinese)
[7] DUBOIS L, ANDERSSON K, ASPLUND A, et al. Evaluating real-time immunohistochemistry on multiple tissue samples, multiple targets and multiple antibody labeling methods[J]. BMC Research Notes, 2013 18(6): 542.
[8] TAMURA K, INOUE K, TAKAHASHI M, et al. Dose-response involvement of constitutive androstane receptor in mouse liver hypertrophy induced by triazole fungicides[J]. Toxicology Letters, 2013, 221(1): 47-56.
[9] 余杏娟,王博,许静.不同抗原修复方法在免疫组化染色的应用比较[J].国际医药卫生导报,2009, 15(11): 1-3.
YU Xingjuan, WANG Bo, XU Jing. Comparison of the effects of different antigen retrieval methods on immuno-histochemical staining[J]. International Medicine and Health Guidance News, 2009, 15(11):1-3.(in Chinese)
[10] 鞠学萍.不同的抗原修复方法在免疫组化染色中的应用[J].中国卫生标准管理,2015,6(11):174-175.
JU Xueping. Application of different antigen retrieval methods in immunohistochemical staining[J]. China Health Standard Management, 2015, 6(11):174-175.(in Chinese)
[11] 邹亮.免疫组化中抗原修复方法及修复液的比较[J].健康研究,2013,33(3):176-178.
ZOU Liang. Antigen retrieval techniques in immunohistochemistry: a comparison of different methods[J]. Health Research, 2013, 33(3): 176-178.(in Chinese)
[12] 骆新兰,林兴滔,罗东兰,等.pH 9.0不同成分的抗原修复液对免疫组织化学染色结果的影响[J].中华病理学杂志,2012,41(3):192-194.
LUO Xinlan, LIN Xingtao, LUO Donglan, et al. Influence of antigen retrieval solution with different compositions in immunohistochemical staining[J]. Chinese Journal of Pathology, 2012, 41(3): 192-194.(in Chinese)
[13] 王琦,王一鸣,刘铁,等.比较不同抗原修复液对免疫组化结果的影响[J].中国实验诊断学,2015(8):1246-1248.
WANG Qi, WANG Yiming, LIU Tie, et al. Comparison of the effects of different antigen retrieval solution on immunohistochemical staining[J]. Chinese Journal of Laboratory Diagnosis, 2015(8): 1246-1248.(in Chinese)
[14] 陈薇,郭爱桃,李亚卓,等.高压抗原修复液的pH值对淋巴组织免疫组化染色的影响[J].海南医学,2015, 26(9):1370-1372.
CHEN Wei, GUO Aitao, LI Yazhuo, et al. Effects of different pH values of high-pressure antigen retrieval buffers on immunohistochemical staining of lymphoid tissue[J]. Hainan Medical Journal, 2015, 26(9): 1370-1372.(in Chinese)
[15] PHANOMSRI E, WARAASAWAPATI S, KHUNMEE S. Multiple immunofluorescence labeling of the same section of invasive ductal breast cancer: a preliminary study[J]. Journal of Medical Technology and Physical Therapy, 2009, 21(2): 104-112.
[16] XIAO Guobing, PAN Cuibao, CAI Yaozhang, et al. Effect of benzene, toluene, xylene on the semen quality and the function of accessory gonad of exposed workers[J]. Industrial Health, 2001, 39(2): 206-210.
[17] RAJAN ST, MALATHI N. Health hazards of xylene: a literature review[J]. Journal of Clinical and Diagnostic Research: JCDR, 2014, 8(2): 271-274.
[18] 柴利,张翠薇,张旭.TO 型生物制片透明剂与二甲苯在病理制片中的应用比较[J].现代医药卫生,2012,28(12):1808-1809.
CHAI Li, ZHANG Cuiwei, ZHANG Xu. Comparison on application of TO type transparent reagent and xylene in pathological section preparation[J]. Modern Medicine & Health, 2012, 28(12): 1808-1809.(in Chinese)


 XU Te,WANG Yun,LI Yanpeng,et al.Identification of biomarkers in human colorectal cancer[J].Journal of Shenzhen University Science and Engineering,2018,35(5):536.[doi:10.3724/SP.J.1249.2018.05536]


Foundation:Openning Project of Key Laboratory of Shaanxi Province for Craniofacial Precision Medicine Research (2018LHM-KFKT007); Shenzhen Science and Technology Research Foundation (JCYJ20160520174247964)
Corresponding author:Professor WANG Yun. E-mail: yunw@szu.edu.cn
Citation:XU Te, WANG Yun, DENG Fang, et al. Technique improvement of multiple immunohistochemical staining on tissue microarray[J]. Journal of Shenzhen University Science and Engineering, 2019, 36(5): 570-575.(in Chinese)
作者简介:许特( 1988—) ,深圳大学博士后研究人员.研究方向: 慢病早期诊断与防治.E-mail:xute@szu.edu.cn
引文:许特,王筠,邓芳,等.组织芯片多重免疫组织化学染色方法的优化[J]. 深圳大学学报理工版,2019,36(5):570-575.
更新日期/Last Update: 2019-09-30