Final Report Summary - PBC-BS (In vivo study of primary B cell responses) The development of a protective immune response to infection is the result of a complex process of activation and differentiation of immune cells that is initiated when a foreign antigen is recognised for a specific lymphocyte. Key aspects of the immune response involve the intrinsic motility of the immune cells themselves, external factors that regulate their trafficking and cell-cell interactions. The process by which immune cells transit through organised lymphoid tissues is crucial to the immune response; however, this issue remains poorly understood. The principal goal of this project was to understand the different factors that modulate lymphocyte activation in vivo and therefore the outcome of an immune response. By taking advantage of new microscopy techniques we have visualised how lymphocytes become activated in living lymphoid organs. This knowledge will allow the design of more specific and efficient vaccination strategies for the treatment of infectious diseases and cancer. Antibodies have demonstrated their value as bio-therapeutics across a wide spectrum of diseases, such as cancer, immune disorders and infection. Thus, understanding the signals required to effectively activate B cells is key for the successful generation of highly specific antibodies. Previous work from our lab and others has established the ability of a special subset of T cells, called NKT cells, to help B cell activation. NKT cells are activated in response to self or foreign antigenic lipids. We have developed a system that allows us to follow the arrival lipids to the lymph nodes, where they are retained within minutes of injection. B cells internalize lipids in the lymph node periphery before polarising to the deeper parts of the nodes where they interact with NKT cells, forming B cell-NKT cell conjugates that are stable for several minutes. These interactions are translated in full B cell activation, proliferation and antibody secretion by antigen-specific B cells. Our results identify B cell internalisation of lipids as a means of modulating NKT-mediated B cell responses in vivo and emphasise the importance of NKT cells in coordinating innate and adaptive immune reactions. NKT cells are master regulators of immunity and their activation has important implications for the outcome of immune responses. Indeed, mice lacking NKT cells have a reduced survival after infection with several pathogens, such as flu virus, Streptococcus pneumoniae and Borrelia. However, the mechanism by which NKT cells encounter antigen and are activated in vivo was not understood. We have identified lymph-node resident macrophages located in the lymph node periphery as key players in presenting antigen to NKT cells. These macrophages are able to retain antigen in the lymph nodes and to efficiently present it to NKT cells inducing their fast activation, proliferation and cytokine secretion. Activation of NKT cells lead to a local response in the lymph node with the recruitment of several types of immune cells including B cells, T cells, dendritic cells and NK cells. We have sketched a comprehensive picture of the cellular interactions that take place in the lymph nodes at the beginning of immune responses. Our findings offer a new perspective on the ways in which the body combats infection and point the way to therapies to treat many common illnesses. Our results indicate that lymph node macrophages can act as a platform to allow antigen presentation to NKT cells at the beginning of an immune response, whereas later B cell-NKT cell interactions will favour the development of adaptive immune responses. The adjuvant-like features of NKT cells make them excellent candidates for their use in vaccination and targeting of specific antigen to lymph node macrophages may allow faster initiation of immune responses.