A neutralizing recombinant human antibody Fab fragment against Puumala hantavirus

A combinatorial human antibody Fab pComb3H library, generated from splenic lymphocytes of a Puumala hantavirus (PUUV) immune individual, was selected against PUUV using the phage display technique. Fanning was carried out wit h antigens immobilized by MAbs directed to the two PUUV envelope glycoprote ins G1 and G2. Thirteen Fabs, with reactivity directed to PUUV and specific ally the G2 protein, as assessed by immunofluorescence and ELISA respective ly, were isolated in crude preparations. By a focus reduction neutralizatio n test (FRNT), four of the 13 crude Fab preparations exhibited type-specifi c neutralization of PUUV (strain Sotkamo) with 44-54% reduction in the numb er of foci. After affinity purification, the four Fab clones exhibited 50% focus reduction of PUUV at concentrations below 2 mu g/ml. Sequencing of th e heavy and light chain complementarity determining regions (CDR) 1-3 showe d that the four selected clones were identical within the antibody binding regions. In inhibition tests with the PUUV G2-specific MAbs, 4G2 and 1C9, a new epitope important for neutralization, designated as G2-a3, was defined . This epitope, overlapping partially the neutralizing epitope recognized b y the human MAb 1C9, seems to be unique for the PUUV serotype since none of the Fab clones neutralized any of the other hantaviruses tested. J. Med. V irol. 60:446-454, 2000. (C) 2000 Wiley-Liss, Inc.