HIF1α特异性人源喉癌噬菌体单链抗体的制备及其对放疗增敏的实验研究

周寒静; 刘静姝; 隆姝孜; 唐翠萍; 张涛

doi:10.16016/j.1000-5404.201612056

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HIF1α特异性人源喉癌噬菌体单链抗体的制备及其对放疗增敏的实验研究

基础医学 | 更新时间：2026-01-09

- HIF1α特异性人源喉癌噬菌体单链抗体的制备及其对放疗增敏的实验研究
- Preparation of human phage displayed single-chain antibody against hypoxia-inducible factor 1 alpha of laryngeal carcinoma and its effect on radiotherapy sensitization
- 第三军医大学学报 2017年39卷第16期页码：1625-1630
- 作者机构：
- 作者简介：
- 基金信息：
  
  重庆市卫生和计生委中医药科技项目(ZY20150245);肿瘤科国家临床重点专科建设项目(国卫办医函(2013)544号)
- DOI：10.16016/j.1000-5404.201612056
  中图分类号：
- 收稿：2016-12-08，
  
  纸质出版：2017-08-30
- 稿件说明：
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周寒静, 刘静姝, 隆姝孜, 等. HIF1α特异性人源喉癌噬菌体单链抗体的制备及其对放疗增敏的实验研究[J]. 第三军医大学学报, 2017,39(16):1625-1630.

Hanjing ZHOU, Jingshu LIU, Shuzi LONG, et al. Preparation of human phage displayed single-chain antibody against hypoxia-inducible factor 1 alpha of laryngeal carcinoma and its effect on radiotherapy sensitization[J]. Journal of Third Military Medical University, 2017, 39(16): 1625-1630.
周寒静, 刘静姝, 隆姝孜, 等. HIF1α特异性人源喉癌噬菌体单链抗体的制备及其对放疗增敏的实验研究[J]. 第三军医大学学报, 2017,39(16):1625-1630. DOI： 10.16016/j.1000-5404.201612056.

Hanjing ZHOU, Jingshu LIU, Shuzi LONG, et al. Preparation of human phage displayed single-chain antibody against hypoxia-inducible factor 1 alpha of laryngeal carcinoma and its effect on radiotherapy sensitization[J]. Journal of Third Military Medical University, 2017, 39(16): 1625-1630. DOI： 10.16016/j.1000-5404.201612056.

摘要

Abstract

Objective

To construct a phage-displayed library and identify phage-displayed single-chain variable fragment (scFv) antibody against hypoxia-inducible factor 1 alpha (HIF1α) of laryngeal carcinoma.

Methods

Total RNA of positive lymph node tissues adjacent to laryngeal carcinoma was extracted. ScFv gene fragments were amplified by RT-PCR and splicing-overlap-extension

(SOE) PCR

then linked into the pCANTAB5E phagemid vector. The recombinant plasmids were transformed into

E.coli

TG1 cells in order to create the primary phage-displayed scFv library. The primary scFv library was performed by biopanning against HEP2 cells and HIF1α antigen. The properties of the product were identified by Western blotting

and its specificity was detected by ELISA and immunocytochemical assay. CCK8 assay was used to detect its effect on cell viability

and cell clone assay was used to detect its effect on survival fraction.

Results

A phage-displayed scFv library targeting HIF1α antigen and HEP2 cells was successfully constructed. The prepared scFv antibody had an apparent molecular weight of 34×10

and down-regulated the expression of HIF1α protein in HEP2 cells. ELISA showed that the obtained scFv antibody had a positive recognition rate of 79% to HIF1α antigen. Immunocytochemical assay indicated that it could specifically bind to HEP2 cells. The obtained scFv antibody decreased the viability of HEP2 cells after X-ray radiation when compared with the cells treated by radiation alone (

0.05). Colony formation test showed that the sensitization enhancement ratio (SER) of the antibody to radiation was 1.89.

Conclusion

A human anti-HIF1α scFv antibody of laryngeal carcinoma is successfully prepared

and it can increase the radiation sensitivity of HEP2 cells.

关键词

Keywords

references

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