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

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.