Construction and development of a mammalian cell-based full-length antibody display library for targeting
Feng Li1, Yan-Hong Liu, Yan-Wen Li
1Cancer Research Institute, Central South University, 110 Xiangya Road, Changsha, 410078, Hunan Province, China.
Abstract:
We present a detailed method for constructing a mammalian cell-based full-length antibody display library for targeting hepatocellular carcinoma. Two novel mammalian library vectors pcDNA3-CHm and pcDNA3-CLm were constructed that contained restriction enzyme sites NheI, ClaI and antibody constant domain. Mammalian expression vector pcDNA3-CHm contains IgG heavy-chain (HC) constant region and glycosylphosphatidylinositol anchor (GPI) that could be anchored full-length antibodies on the surface of mammalian cells. GOLPH2 prokaryotic expression vector was carried out in Escherichia coli and purified by immobilized metal affinity chromatography. Variable domain of heavy-chain and variable domain of light-chain genes were respectively inserted into the vector pcDNA3-CHm and pcDNA3-CLm by ligation, and antibody libraries are displayed as whole IgG molecules on the cell surface by co-transfecting this HC-GPI with a light chain. By screening the cell library using magnetic beads and cell ELISA, the cell clone that displayed GOLPH2-specific antibodies on cell surfaces was identified. The mammalian cell-based antibody display library is a great potential application for displaying full-length functional antibodies of targeting hepatocellular carcinoma on the surface of mammalian cells. Anti-GOLPH2 display antibody was successfully isolated from the library.
Insights
We developed a mammalian cell display library for hepatocellular carcinoma (HCC) targeting. This method successfully isolated anti-GOLPH2 antibodies, showing potential for HCC therapeutic antibody discovery.
Area of Science:
- Biotechnology
- Immunology
- Oncology
Background:
- Hepatocellular carcinoma (HCC) remains a significant global health challenge.
- Developing targeted therapies requires novel antibody discovery platforms.
- Full-length antibody display on mammalian cells offers advantages for functional screening.
Purpose of the Study:
- To establish a mammalian cell-based full-length antibody display library for hepatocellular carcinoma (HCC) targeting.
- To construct novel mammalian expression vectors for cell surface antibody display.
- To screen the library for antibodies targeting HCC-associated antigens.
Main Methods:
- Construction of novel mammalian expression vectors (pcDNA3-CHm, pcDNA3-CLm) encoding antibody constant regions and glycosylphosphatidylinositol (GPI) anchor.
- Insertion of variable heavy-chain and variable light-chain genes into display vectors.
- Co-transfection of vectors for cell surface display of full-length IgG antibodies.
- Screening of the antibody display library using magnetic beads and cell ELISA.
Main Results:
- Successful construction of a mammalian cell-based full-length antibody display library.
- Identification and isolation of a cell clone displaying GOLPH2-specific antibodies.
- Demonstration of the platform's capability to display functional antibodies on mammalian cell surfaces.
Conclusions:
- The developed mammalian cell-based antibody display library is a promising platform for discovering functional antibodies against hepatocellular carcinoma.
- The isolation of anti-GOLPH2 antibodies highlights the potential of this system for identifying targeted cancer therapies.
- This methodology facilitates the display of full-length antibodies on cell surfaces for various therapeutic applications.
