ABSTRACT – PROJECT 1 Much of the focus of vaccine-induced antibody responses to prevent HIV acquisition has been on IgG, the most common antibody isotype. However, previous work by this HIVRAD has found that combinations of IgA and IgG can inhibit SIV acquisition under circumstances where either alone shows no or limited inhibitory activity. This demonstrates the importance of considering the combined actions of the effector domains of multiple antibody isotypes in the successful developments of a vaccine. To gain mechanistic insights into the underlying ability of these antibody combinations to inhibit rectal acquisition in the rhesus macaque model, we will utilize our toolbox of microscopy-based reagents to gain insights into how acquisition is decreased or blocked. These tools allow us to directly examine the major components of the vaccine virus interaction at the mucosal surface including the labeling of antibodies, virus, and target cells, all in the proper anatomical configuration of the rectal compartment of a living animal. This plan of attack will take advantage of support from the project cores including macaque veterinary support, antibodies and virions, and the ability to do in vivo imaging via PET scan. New antibody combinations will come from data generated by project 2. More complicated vaccine induced responses will come from interactions with project 3. These studies will provide critical insights into the behavior of antibodies and their interactions with virions at mucosal surfaces. For example, we will determine the percentage of IgG at the rectal mucosa that is either generated systemically or locally. We will also determine the stability of antibodies within mucosal tissues. Understanding how antibody virus interactions influence the localization and phenotype of infected cells will further advance our knowledge of vaccine action. Understanding the in vivo mechanisms of IgA action against SHIV will also inform new possibilities in vaccine design. This information will inform vaccine development across a variety of pathogens. By providing mechanistic insights into the underlying effector functions of antibodies in preventing acquisition, we will advance our ability to develop a vaccine, which will protect humans from HIV acquisition.