title: Construction and development of a mammalian cell-based full-length antibody display library for targeting hepatocellular carcinoma.
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.
authors: Cancer Research Institute, Central South University, 110 Xiangya Road, Changsha, 410078, Hunan Province, China.
authors: Feng
authors: F
authors: Li
authors: Yan-Hong
authors: YH
authors: Liu
authors: Yan-Wen
authors: YW
authors: Li
authors: Yue-Hui
authors: YH
authors: Li
authors: Ping-Li
authors: PL
authors: Xie
authors: Qiang
authors: Q
authors: Ju
authors: Lin
authors: L
authors: Chen
authors: Guan-Cheng
authors: GC
authors: Li
mesh_terms: D000912:Antibodies, Neoplasm / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D006528:Carcinoma, Hepatocellular / Q000276:immunology* / Q000473:pathology*; D002460:Cell Line; D061505:Cell Surface Display Techniques / Q000379:methods; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D007074:Immunoglobulin G / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification; D019151:Peptide Library; D011994:Recombinant Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification
chemical_list: D000912:Antibodies, Neoplasm; D007074:Immunoglobulin G; D019151:Peptide Library; D011994:Recombinant Proteins