title: Cloning and expression of two human recombinant monoclonal Fab fragments specific for EBV viral capsid antigen.
abstract: Serum titers of antibody to Epstein-Barr virus (EBV) viral capsid antigen (VCA) have been positively correlated with malignancies of lymphoid proliferation, such as Burkitt's lymphoma and Hodgkin's lymphoma. We have constructed a phage display combinatorial antibody Fab library from a patient with marginal zone B cell lymphoma associated with Sjögren's syndrome and carrying high serum anti-EBV-VCA IgG titer. Fab fragments were selected by panning against EBV-VCA protein coated onto ELISA plates, and selected Fab clones were characterized by ELISA, western blotting (WB), indirect immunofluorescence assay and immunohistochemistry. We have established two Fab clones, Fab-aVCA1 and Fab-aVCA21, which specifically recognize EBV-VCA by ELISA and WB. Inhibition ELISA competition showed that both clones could significantly reduce the binding of specific anti-EBV-VCA mAb to its relevant proteins. Furthermore, these two Fab clones could localize VCA protein in the EBV-positive P3HR1 and Daudi cell lines, as well as in tissue samples from patients with EBV-infected lymphoid malignancies. These results indicate that our two Fab clones are novel human mAbs specific for EBV-VCA protein and may have potential benefits for development of novel diagnostic and therapeutic approaches in EBV-related lymphoid malignancies.
authors.affiliation: Department of Hematology and Immunology, Kanazawa Medical University, Uchinada, Ishikawa 920-0293, Japan.
authors.forename: Lingli
authors.initials: L
authors.lastname: Dong
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Masaki
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Takegami
authors.forename: Takafumi
authors.initials: T
authors.lastname: Kawanami
authors.forename: Kunihiko
authors.initials: K
authors.lastname: Itoh
authors.forename: Zhe-Xiong
authors.initials: ZX
authors.lastname: Jin
authors.forename: Cheng-Ri
authors.initials: CR
authors.lastname: Huang
authors.forename: Xiao-Peng
authors.initials: XP
authors.lastname: Tong
authors.forename: Toshihiro
authors.initials: T
authors.lastname: Fukushima
authors.forename: Masao
authors.initials: M
authors.lastname: Tanaka
authors.forename: Toshioki
authors.initials: T
authors.lastname: Sawaki
authors.forename: Tomoyuki
authors.initials: T
authors.lastname: Sakai
authors.forename: Susumu
authors.initials: S
authors.lastname: Sugai
authors.forename: Toshiro
authors.initials: T
authors.lastname: Okazaki
authors.forename: Yuko
authors.initials: Y
authors.lastname: Hirose
authors.forename: Hisanori
authors.initials: H
authors.lastname: Umehara
mesh_terms: D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D000914:Antibodies, Viral / Q000097:blood*; D000956:Antigens, Viral / Q000276:immunology*; D015153:Blotting, Western; D036022:Capsid Proteins / Q000276:immunology*; D002460:Cell Line; D003001:Cloning, Molecular; D020650:Combinatorial Chemistry Techniques; D004797:Enzyme-Linked Immunosorbent Assay; D020031:Epstein-Barr Virus Infections / Q000150:complications / Q000276:immunology*; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D016393:Lymphoma, B-Cell / Q000209:etiology / Q000276:immunology* / Q000821:virology; D008969:Molecular Sequence Data; D019151:Peptide Library; D020539:Sequence Analysis, Protein; D012859:Sjogren's Syndrome / Q000150:complications* / Q000276:immunology
chemical_list: D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; D000956:Antigens, Viral; D036022:Capsid Proteins; C041788:Epstein-Barr viral capsid antigen; D007140:Immunoglobulin Fab Fragments; D019151:Peptide Library