PROJECT SUMMARY/ABSTRACT Broadly neutralizing antibodies (bNAbs) are likely to be an important component of an HIV-1 vaccine. The HIV- 1 Envelope (Env) glycoprotein is the sole target of bNAbs and has been a focus of immunogen designs. Yet diverse recombinant (rec) Envs do not display detectable binding to the inferred germline (gl) of certain bNAbs, such as the VRC01-class bNAbs, which are the focus of this proposal. Rec Envs capable of binding glVRC01 bNAbs have been designed and induced the production of glVRC01 bNAbs when used as immunogens. However, these novel immunogens are not sufficient to induce the maturation of glVRC01-class bNAbs to their broad neutralization forms. Here we propose to use novel reagents as immunogens to guide the development of VRC01-class bNAbs by: 1) immunizing knock-in mice with immunogens harboring short glycans to evaluate if these could more efficiently prime immune responses that will accommodate the HIV-1 Env glycan shield and 2) combining immunological and biophysical (including atomic level structures) characterization. We hypothesize that such immunogens will already know how to accommodate short glycans and thus be more amenable to maturation. Our preliminary data show that such antigen has increased binding to precursors glVRC01-like antibodies. The sequencing of monoclonal antibodies (mAbs) along with structural information will provide molecular details on the interactions between the vaccine-elicited antibodies from animals and the immunogens: better understanding the epitopes targeted by the novel vaccine-elicited mAbs will help us identify the best boost immunogen for future immunizations studies and thus guide HIV-1 vaccine design.