Identification of Two Competition‐Defined Neutralizing Antibody Groups Targeting the HHV‐6B gH/gL/gQ1/gQ2 Complex Independent of CD134 Binding

ABSTRACT Human herpesvirus 6B (HHV‐6B) entry is mediated by the interaction between gH/gL/gQ1/gQ2 complex (tetramer) and CD134 expressed on activated T cells. In this study, we identified and characterized a panel of monoclonal antibodies targeting the HHV‐6B tetramer, with particular focus on three antibodies; gHgL J1, gHgL O2, and gHgL F5; that represent anti‐gH neutralizing monoclonal antibodies with binding characteristics distinct from the previously described antibody OHV‐3. Immunofluorescence and Western blot analyses confirmed recognition of the HHV‐6B tetramer in infected and transfected cells. Biolayer interferometry (BLI) demonstrated that these antibodies exhibited dissociation constants ( K D ) in the range of 62.52–76.99 nM. We next assessed the effect of antibodies on the interaction between the HHV‐6B tetramer and CD134. None of the gH‐targeting antibodies inhibited the CD134 binding. Competition analysis using BLI‐based antibody‐Fab competition assays indicated that gHgL J1, gHgL O2, and gHgL F5 form a competition‐defined antigenic cluster distinct from that recognized by OHV‐3. Collectively, these findings identify a group of anti‐gH antibodies which neutralizes HHV‐6B with CD134‐binding‐independent profiles and suggest the presence of at least two distinguishable antibody‐recognition regions within HHV‐6B gH. These results expand our understanding of the antigenic organization of the HHV‐6B entry complex.