Targeted detecting HER2 expression with recombinant anti HER2 ScFv-GFP fusion antibody

科研成果详情

题名	Targeted detecting HER2 expression with recombinant anti HER2 ScFv-GFP fusion antibody
作者	Guohui Gao; Chong Chen; Yanmei Yang; Han Yang; Jindan Wang; Yi Zheng; Qidi Huang; Xiaoqu Hu
发表日期	2012-08-01
发表期刊	Sheng wu gong cheng xue bao = Chinese journal of biotechnology 影响因子和分区
语种	英语
原始文献类型	Article
关键词	Anti HER2 single-chain Fv antibody Clinical HER2 detection function Fusion antibody Green fluorescent protein HER2 positive cancer cells Molecular diagnostics
摘要	To verify the reliability of targeted detecting HER2 positive cancer cells and clinical pathological tissue specimens with a recombinant anti HER2 single chain antibody in single chain Fv fragment (scFv) format, we have constructed the fusion variable regions of the ScFv specific for HER2/neu. labeled a green-fluorescent protein(GFP). The humanized recombinant Anti HER2 ScFv-GFP gene was inserted into pFast Bac HT A, and expressed in insect cells sf9. Then the recombinant fusion protein Anti HER2 ScFv-GFP was properly purified with Ni2+-NTA affinity chromatography from the infected sf9 cells used to test the specificity of the fusion antibody for HER2 positive cancer cells. Firstly, the purified antibody incubated with HER2 positive breast cancer cells SKBR3, BT474 and HER2 negative breast cancer cells MCF7 for 12 h/24 h/48 h at 37 degrees C, in order to confirm targeted detecting HER2 positive breast cancer cells by Laser Confocal Microscopy. Furthermore, the same clinical pathological tissue samples were assessed by immunohistochemistry (IHC) and the fusion antibody Anti HER2 ScFv-GFP in the meanwhile. The data obtained indicated that the recombinant eukaryotic expression plasmid pFast Bac HT A/Anti HER2 ScFv-GFP was constructed successfully In addition, obvious green fluorescent was observed in insect cells sf9. When the purified fusion antibody was incubated with different cancer cells, much more green fluorescent was observed on the surface of the HER2 positive cancer cells SKBR3 and BT474. In contrast, no green fluorescent on the surface of the HER2 negative cancer cells MCF7 was detected. The concentration of the purified fusion antibody was 115.5 microg/mL, of which protein relative molecular weight was 60 kDa. The analysis showed the purity was about 97% and the titer was about 1:64. The detection results of IHC and fusion antibody testing indicated the conformity. In summary, the study showed that the new fusion antibody Anti HER2 ScFv-GFP can test HER2 positive cancer cells, indicating a potential candidate method for clinical HER2 positive specimens detection.
ISSN	1000-3061
卷号	28 期号:8 页码:1002-14.
收录类别	PUBMED ; SCOPUS
URL	查看原文
PubMed ID	23185900
SCOPUSEID	2-s2.0-84867623070
文献类型	期刊论文
条目标识符	https://kms.wmu.edu.cn/handle/3ETUA0LF/63541
专题	检验医学院（生命科学学院、生物学实验教学中心）
作者单位	Zhejiang Provincial Key Laboratory of Medical Genetics, School of Life Sciences, Wenzhou Medical College, Wenzhou 325035, Zhejiang, China.
第一作者单位	检验医学院（生命科学学院、生物学实验教学中心）
第一作者的第一单位	检验医学院（生命科学学院、生物学实验教学中心）
推荐引用方式 GB/T 7714	Guohui Gao,Chong Chen,Yanmei Yang,et al. Targeted detecting HER2 expression with recombinant anti HER2 ScFv-GFP fusion antibody[J]. Sheng wu gong cheng xue bao = Chinese journal of biotechnology,2012,28(8):1002-14..
APA	Guohui Gao., Chong Chen., Yanmei Yang., Han Yang., Jindan Wang., ... & Xiaoqu Hu. (2012). Targeted detecting HER2 expression with recombinant anti HER2 ScFv-GFP fusion antibody. Sheng wu gong cheng xue bao = Chinese journal of biotechnology, 28(8), 1002-14..
MLA	Guohui Gao,et al."Targeted detecting HER2 expression with recombinant anti HER2 ScFv-GFP fusion antibody".Sheng wu gong cheng xue bao = Chinese journal of biotechnology 28.8(2012):1002-14..