Vaccines are one of the greatest achievements to public health allowing almost complete eradication of small pox and polio and dramatically reducing the incidence of a variety of other diseases. However, many diseases still exist without a vaccine. To create vaccines for these diseases we need to understand and create enhanced responses that enable proper immune activation. This organized activation can be achieved using adjuvants, components added to the vaccine to enhance the immune response. Typically, vaccines have been empirically derived, leading to expensive and lengthy development periods. Our lab is focused on creating tools to enable rational and optimized vaccine design. My work focuses on two main areas: creating tools to probe immune responses on the single-cell level, leading to a greater understanding of immune activation and applying this understanding to create more effective vaccines to challenging diseases. One of the biggest challenges of creating new vaccines is attaining an adequate safety profile. Many vaccines can provide protection but do not translate to the clinic due to the high levels of inflammation they induce. CpG, a synthetic bacterial DNA mimic, has demonstrated great promise as an adjuvant, however most vaccines that include CpG do not make it through clinical trials. Using an NF-kB inhibitor, we demonstrate that we can enhance the safety and protection afforded by CpG and many other common adjuvants. Many of the most effective vaccines stimulate multiple innate immune pathways. When this combination of pathways leads to enhancement of the immune response this is known as an immune synergy. Although the existence of immune synergies is well understood, the mechanism of enhanced activity is still unknown. Few tools exist to directly examine spatial and temporal elements of immune activation and synergies. Described here are two tools to elucidate the spatiotemporal aspects of innate immune responses. The first is a particle-based system allowing effective agonist presentation and tracking of activated cells. The second is an optogenetic innate immune receptor allowing the receptor to be activated with the spatial and temporal precision of light.