Monoclonal antibody preparation of Golgi phosphoprotein 2 and preliminary application in the early diagnosis of hepatocellular carcinoma

Ju,Qiang; Zhao,Yanjie; Liu,Yanhong; Zhou,Guohua; Li,Feng; Xie,Pingli; Li,Yuehui; Li,Guan-Cheng

doi:10.3892/mmr.2013.1503

August 2013 Volume 8 Issue 2

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August 2013 Volume 8 Issue 2

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Article

Monoclonal antibody preparation of Golgi phosphoprotein 2 and preliminary application in the early diagnosis of hepatocellular carcinoma

Authors:
- Qiang Ju
- Yanjie Zhao
- Yanhong Liu
- Guohua Zhou
- Feng Li
- Pingli Xie
- Yuehui Li
- Guan-Cheng Li
View Affiliations / Copyright

Affiliations: Cancer Research Institute, Key Laboratory of Carcinogenesis and Cancer Invasion Ministry of Education, Key Laboratory of Carcinogenesis Ministry of Health, Central South University, Changsha, Hunan 410078, P.R. China
Pages: 517-522
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Published online on: May 31, 2013

https://doi.org/10.3892/mmr.2013.1503
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Abstract

Golgi phosphoprotein 2 (Golph2) is a type II Golgi‑specific membrane protein, which has been found to be overexpressed in hepatocellular carcinoma (HCC) patients. The sensitivity of diagnosis of HCC using Golph2 (76%) was markedly elevated compared with alpha‑fetoprotein (AFP) (70%), and Golph2 is expected to be a novel and effective serum biomarker for the diagnosis of HCC. The aim of this study was to prepare monoclonal antibodies against Golph2 and to establish double-antibody sandwich enzyme-linked immunosorbent assay (s-ELISA), which will be used in diagnostics, therapeutics and as a tool in understanding the role of Golph2 in the pathogenesis of liver diseases and cancer. In this study, fusion protein TRX-Golph2 was expressed and purified using an Escherichia coli system. BALB/c mice were immunized with TRX-Golph2 recombinant protein. The hybridoma technique was used for the production of anti-Golph2 monoclonal antibody. Hybridoma clones were screened using indirect ELISA and anti-Golph2 monoclonal antibody was produced in the ascites of BALB/c mice. The specificity of anti-Golph2 monoclonal antibody was detected by western blot analysis and immunocytochemistry. s-ELISA was established using horseradish peroxidase (HRP)‑labeled anti-Golph2 monoclonal antibody and used to detect the antigen in the serum of HCC patients. As a result, five stable hybridoma cell clones (5C6D5, 5B7F5, 7F5F3, 8A7B4, 8C9E8) producing anti-Golph2 monoclonal antibody were established. The highest titer of anti-Golph2 monoclonal antibody (5C6D5) was 1:51,200. Western blot analysis revealed that anti-Golph2 monoclonal antibody had a high specificity for Golph2 protein. Anti-Golph2 monoclonal antibody was HRP-labeled and the optimal working concentration was found to be 1:500. The levels of antigen in a proportion of HCC patients were shown to be significantly higher compared to those found in healthy controls.

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Journals

International Journal of Molecular Medicine

International Journal of Oncology

Molecular Medicine Reports

Oncology Reports

Experimental and Therapeutic Medicine

Oncology Letters

Biomedical Reports

Molecular and Clinical Oncology

World Academy of Sciences Journal

International Journal of Functional Nutrition

International Journal of Epigenetics

Medicine International

Monoclonal antibody preparation of Golgi phosphoprotein 2 and preliminary application in the early diagnosis of hepatocellular carcinoma

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