Prokaryotic Expression of Anti-TAG-72 Single-Chain Variable Fragment and Its Value in Detection of Hepatocellular Carcinoma_CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Authors：

XU Hongyong ¹ , XU Li ² , LI Kaizong ³ , DOU Kefeng ³

1.Department of Hepatobiliary General Surgery, The 451st Hospital of Chinese PLA, Xi’an 710054, China;;
2.Institute of Digestive Disease, Xijing Hospital, The Fourth Medical University, Xi’an 710032, China;;
3.Department of Hepatobiliary Surgery, Xijing Hospital, The Fourth Medical University, Xi’an 710032, China;

Corresponding author：

Keywords：

Hepatocellular carcinoma; Single-chain fragment variable antibody; Prokaryotic expression; Tumor associated glycoprotein-72 antigen

Video：

Export PDF Favorites Scan Get Citation

Abstract Full text Figures/Tables Video References Cited by

Objective To study the prokaryotic expression of the anti-tumor associated glycoprotein-72 (TAG-72) singlechain fragment variable (scFv) antibody and its specific affinity to hepatocellular carcinoma cell lines and tissues. Methods The cDNA of anti-TAG-72 scFv antibody was inserted into pCANTAB5E to obtain phage vector anti-TAG-72 -scFv-pCANTAB5E. Isopropyl-β-D-thiogalactoside (IPTG) was used to induce the expression of anti-TAG-72 scFv antibody. SDS-PAGE and Western blot were used to identify the anti-TAG-72 scFv antibody. Human hepatocellular carcinoma cells were cultured, and TAG-72 was determined with the obtained scFv by immunohistochemistry in the cells and paraffin-embedded hepatocellular carcinoma tissues.
Results SDS-PAGE and Western blot showed that the anti-TAG-72-scFv antibody was successfully expressed. Anti-TAG-72 scFv could bind to hepatocellular carcinoma cell lines SMMC7721, HepG2, and HHCC, but not to BEL7402, suggesting that SMMC7721, HepG2, and HHCC cells expressed TAG-72. For the 40 cases of hepatocellular carcinoma tissues, the positive rate of TAG-72 in stage Ⅰ and Ⅱ-Ⅲ was 23.08% (3/13) and 62.96%(17/27), respectively. While no TAG-72 expression was found in the 10 normal cases. TAG-72 expression was significantly different between hepatocellular carcinomas and normal tissues (P lt;0.05).
Conclusions The prokaryotic expression of anti-TAG-72 scFv antibody is successfully achieved, and can be used to identify TAG-72 antigen. TAG-72 is highly expressed in hepatocellular carcinoma, but not in normal liver tissue, which may be suggested as cancer marker of hepatocellular carcinoma.

Citation： XU Hongyong ,XU Li,LI Kaizong,DOU Kefeng. Prokaryotic Expression of Anti-TAG-72 Single-Chain Variable Fragment and Its Value in Detection of Hepatocellular Carcinoma. CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY, 2010, 17(7): 699-702. doi: Copy

1.	Xu M, Real FX, Welt S, et al. Expression of TAG72 in normal colon, transitional mucosa, and colon cancer [J]. Int J Cancer, 1989； 44(6): 　985989.
2.	Friedman E, Thor A, Hand PH, et al. Surface expression of tumorassociated antigens in primary cultured human colonic epithelial cells from carcinomas, benign tumors, and normal tissues [J]. Cancer Res, 1985； 45(10): 　 56485655.
3.	Thor A, Ohuchi N, Szpak CA, et al. Distribution of oncofetal antigen tumorassociated glycoprotein72 defined by monoclonal antibody B72.3 [J]. Cancer Res, 1986; 46(6): 　 31183124.
4.	徐宏勇, 徐立, 李开宗，等. 抗肿瘤相关糖蛋白72单抗在人乳腺癌中的表达 [J]. 中国普外基础与临床杂志, 2006; 13(5):　 540541, 549.
5.	徐宏勇, 徐立, 李开宗, 等. 靶向性肿瘤相关抗原TAG72的scFvCD28嵌合基因的真核表达载体的构建 [J]. 中国普外基础与临床杂志, 2002; 9(6): 　 398401.
6.	徐宏勇, 徐立, 高建宏, 等. 肿瘤相关糖蛋白72抗原在胆囊癌中的表达及其临床意义[J]. 山东医药， 2007; 47(3): 　 1618.
7.	金冬雁, 黎孟枫. 分子克隆实验指南 [M]. 第2版. 北京: 　科学出版社, 2002: 　 4956.
8.	梁英杰, 黎红, 赖英荣. 免疫组织化学LDP染色法在病理诊断中的应用 [J]. 中华病理学杂志， 1999; 28(1): 　 6061.
9.	Ponnusamy MP, Venkatraman G, Singh AP, et al. Expression of TAG72 in ovarian cancer and its correlation with tumor stage and patient prognosis [J]. Cancer Lett， 2007； 251(2): 　247257.
10.	Chauhan SC, Vinayek N, Maher DM, et al. Combined staining of TAG72, MUC1, and CA125 improves labeling sensitivity in ovarian cancer: 　 antigens for multitargeted antibodyguided therapy [J]. J Histochem Cytochem， 2007; 55(8): 　867875.
11.	徐宏勇, 徐立, 李开宗, 等. 癌胚抗原嵌合锚定T细胞对胃癌的体内外抑癌效应观察 [J]. 中华外科杂志, 2008; 46(2): 　 148149.
12.	徐宏勇, 徐立, 高建宏, 等. 嵌合锚定T细胞致癌胚抗原阳性胃癌细胞的凋亡效应 [J]. 中华医学杂志, 2007; 87（15）: 　 10531057.

1. Xu M, Real FX, Welt S, et al. Expression of TAG72 in normal colon, transitional mucosa, and colon cancer [J]. Int J Cancer, 1989； 44(6): 　985989.
2. Friedman E, Thor A, Hand PH, et al. Surface expression of tumorassociated antigens in primary cultured human colonic epithelial cells from carcinomas, benign tumors, and normal tissues [J]. Cancer Res, 1985； 45(10): 　 56485655.
3. Thor A, Ohuchi N, Szpak CA, et al. Distribution of oncofetal antigen tumorassociated glycoprotein72 defined by monoclonal antibody B72.3 [J]. Cancer Res, 1986; 46(6): 　 31183124.
4. 徐宏勇, 徐立, 李开宗，等. 抗肿瘤相关糖蛋白72单抗在人乳腺癌中的表达 [J]. 中国普外基础与临床杂志, 2006; 13(5):　 540541, 549.
5. 徐宏勇, 徐立, 李开宗, 等. 靶向性肿瘤相关抗原TAG72的scFvCD28嵌合基因的真核表达载体的构建 [J]. 中国普外基础与临床杂志, 2002; 9(6): 　 398401.
6. 徐宏勇, 徐立, 高建宏, 等. 肿瘤相关糖蛋白72抗原在胆囊癌中的表达及其临床意义[J]. 山东医药， 2007; 47(3): 　 1618.
7. 金冬雁, 黎孟枫. 分子克隆实验指南 [M]. 第2版. 北京: 　科学出版社, 2002: 　 4956.
8. 梁英杰, 黎红, 赖英荣. 免疫组织化学LDP染色法在病理诊断中的应用 [J]. 中华病理学杂志， 1999; 28(1): 　 6061.
9. Ponnusamy MP, Venkatraman G, Singh AP, et al. Expression of TAG72 in ovarian cancer and its correlation with tumor stage and patient prognosis [J]. Cancer Lett， 2007； 251(2): 　247257.
10. Chauhan SC, Vinayek N, Maher DM, et al. Combined staining of TAG72, MUC1, and CA125 improves labeling sensitivity in ovarian cancer: 　 antigens for multitargeted antibodyguided therapy [J]. J Histochem Cytochem， 2007; 55(8): 　867875.
11. 徐宏勇, 徐立, 李开宗, 等. 癌胚抗原嵌合锚定T细胞对胃癌的体内外抑癌效应观察 [J]. 中华外科杂志, 2008; 46(2): 　 148149.
12. 徐宏勇, 徐立, 高建宏, 等. 嵌合锚定T细胞致癌胚抗原阳性胃癌细胞的凋亡效应 [J]. 中华医学杂志, 2007; 87（15）: 　 10531057.

CHINESE JOURNAL OF BASES AND CLINICS IN GENERAL SURGERY

Prokaryotic Expression of Anti-TAG-72 Single-Chain Variable Fragment and Its Value in Detection of Hepatocellular Carcinoma

Abstract Full text Figures/Tables Video References Cited by

Format

Content