Isolation of potent neutralizing antibodies from a survivor of the 2014 Ebola virus outbreak

Abstract

Antibodies targeting the Ebola virus surface glycoprotein (EBOV GP) are implicated in protection against lethal disease, but the characteristics of the human antibody response to EBOV GP remain poorly understood. Here we isolated and characterized 349 GP-specific monoclonal antibodies (mAbs) from the peripheral B cells of a convalescent donor who survived the 2014 EBOV Zaire outbreak. Remarkably, 77% of the mAbs neutralize live EBOV and several mAbs exhibit unprecedented potency. Structures of selected mAbs in complex with GP reveal a site of vulnerability located in the GP stalk region proximal to the viral membrane. Neutralizing antibodies (NAbs) targeting this site show potent therapeutic efficacy against lethal EBOV challenge in mice. The results provide a framework for the design of new EBOV vaccine candidates and immunotherapies.

Exploring Human Antimicrobial Antibody Responses on a Single B Cell Level

“…The antimicrobial human antibody repertoire can be studied by single B cell isolation and in-depth repertoire sequencing (Fig. 1); both aspects have proven to be powerful techniques for the characterization of immune responses to pathogen-specific antigens (4,5). Following the isolation of peripheral blood mononuclear cells (PBMCs) from affected individuals, antigen-specific plasmablasts or memory B cells can be sorted by fluorescence-activated cell sorting (FACS) by labeling the cells with a fluorescently tagged antigen or by sorting of naive plasmablasts in acutely infected individuals (Fig.…”

“…After isotype-specific amplification using appropriate antisense primers for the CH1 region of, e.g., IgG, IgM, or IgA, VH and VL regions can be grafted onto the respective constant domains. Subsequently, antibodies can be recombinantly expressed and processed for further characterization (4,6,7). Thus, recombinant MAbs can be immediately obtained without the need for B cell immortalization or fusion with myeloma partners.…”

“…The characterization of single MAbs in complex with the respective antigen provides insights into the identification of novel mechanisms and epitopes required for successful protection against the pathogen. Those MAbs with potent protective ability against a broad range of related pathogens or strains (broadly neutralizing antibodies [bNAbs]) are of great interest for antibody discovery and are promising candidates for immunotherapeutic development (4).…”

“…Rapid development of immunotherapies for newly emerging pathogens, or analysis of antibody repertoires, depends on highly sophisticated technology platforms such as single B cell sorting or hybridoma technology. In acute cases, it is important to utilize the most rapid direct method for the isolation of potent human antibodies to generate therapeutic molecules for clinical applications-even within a few weeks (4). In comparison to hybridoma technology or immunization of transgenic animals (e.g., with humanized Ig loci), there are significant advantages of B cell sorting.…”

“…Murine antibodies were also shown to trigger immunogenic responses, provide less stability, and hamper downstream Fc effector functions. On the basis of all of these pitfalls, our laboratory and those of others use antigen-dependent B cell sorting for the isolation of a variety of different pathogen-specific molecules for a straightforward MAb isolation within a few weeks (4,30). For the detection of antigen-specific B cell populations, the antigen must be labeled either by fluorescently labeled streptavidin or by another fluorescent tag.…”