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International Immunology Advance Access originally published online on January 31, 2007
International Immunology 2007 19(3):331-336; doi:10.1093/intimm/dxl150
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© The Japanese Society for Immunology. 2007. All rights reserved. For permissions, please e-mail: journals.permissions@oxfordjournals.org

Cloning and expression of two human recombinant monoclonal Fab fragments specific for EBV viral capsid antigen

Lingli Dong1,3, Yasufumi Masaki1, Tsutomu Takegami2, Takafumi Kawanami1, Kunihiko Itoh4, Zhe-Xiong Jin1, Cheng-Ri Huang1, Xiao-Peng Tong1, Toshihiro Fukushima1, Masao Tanaka1, Toshioki Sawaki1, Tomoyuki Sakai1, Susumu Sugai1, Toshiro Okazaki5, Yuko Hirose1 and Hisanori Umehara1

1 Department of Hematology and Immunology
2 Department of Molecular Oncology and Virology, Kanazawa Medical University, Uchinada, Ishikawa 920-0293, Japan
3 Department of Hematology and Immunology, Tongji Hospital, Huazhong University of Science and Technology, Wuhan, Hubei 430030, China
4 Department of Clinical Pharmacology and Genetics, School of Pharmaceutical Science, University of Shizuoka, Shizuoka 422-8526, Japan
5 Department of Clinical Laboratory, Medicine/Hematology, Faculty of Medicine, Tottori University, Yonago, Tottori 683-8504, Japan

Correspondence to: Y. Masaki; E-mail: yasum{at}kanazawa-med.ac.jp

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.

Keywords: antibodies, human, molecular biology

Transmitting editor: K. Okumura

Received 4 September 2006, accepted 22 December 2006.


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