title: Generation and characterization of a novel recombinant antibody against LMP1-TES1 of Epstein-Barr virus isolated by phage display.
abstract: Latent Membrane Protein 1 (LMP1) is a primary target for controlling tumorigenesis in Epstein-Barr virus related malignancies; in this study, we aimed to develop a specific antibody against the TES1 domain of the oncogenic LMP1. We screened a full human naïve Fab phage library against TES1 peptide, which consisted of C terminal-activating regions proximal 44 amino acids. After three rounds of panning, enrichment and testing by phage ELISA and further analyzed by DNA sequencing, we selected a phage clone with the highest affinity to LMP1-TES1 and designated it as htesFab. The positive clone was expressed in Escherichia coli and the purified htesFab was characterized for its binding specificity and affinity to LMP1. ELISA, immunofluorescence and FACS analysis confirmed that htesFab could recognize LMP1 TES1 both in vitro and in LMP1 expressing HNE2-LMP1 cells. Furthermore, MTT assay showed that htesFab inhibited the proliferation of HNE2-LMP1 cells in a dose-dependent manner. In summary, this study reported the isolation and characterization of human Fab, which specifically targets the C terminal region/TES1 of LMP1, and has potential to be developed as novel tool for the diagnosis and therapy of Epstein-Barr virus related carcinoma.
authors.affiliation: Department of Otolaryngology Head and Neck Surgery, The Second Affiliated Hospital of Nanjing Medical University, 121 Jiangjiayuan, Nanjing 210011, China. chinarenzdw@yahoo.com.cn
authors.forename: Dawei
authors.initials: D
authors.lastname: Zhang
authors.forename: Yuan
authors.initials: Y
authors.lastname: Mao
authors.forename: Qing
authors.initials: Q
authors.lastname: Cao
authors.forename: Lin
authors.initials: L
authors.lastname: Xiong
authors.forename: Juan
authors.initials: J
authors.lastname: Wen
authors.forename: Renjie
authors.initials: R
authors.lastname: Chen
authors.forename: Jin
authors.initials: J
authors.lastname: Zhu
mesh_terms: D000595:Amino Acid Sequence; D000906:Antibodies / Q000276:immunology*; D002460:Cell Line; D061505:Cell Surface Display Techniques; D004854:Herpesvirus 4, Human / Q000276:immunology*; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D019151:Peptide Library; D011485:Protein Binding / Q000276:immunology; D011993:Recombinant Fusion Proteins / Q000276:immunology* / Q000378:metabolism; D014763:Viral Matrix Proteins / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism
chemical_list: D000906:Antibodies; C037662:EBV-associated membrane antigen, Epstein-Barr virus; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library; D011993:Recombinant Fusion Proteins; D014763:Viral Matrix Proteins