Abstract Recombinant DNA vaccines show incredible promise for the prevention of human disease by their capacity to effectively inducing both humoral and cellular immune responses. Among the available technologies adenovirus (Ad) vectored vaccines are among the most appealing. To date the clinical deployment of Ad-based vaccines has been limited by several considerations including the lack of delivery technology to enable safe, reproducible and effective vector delivery to patients. The skin is readily accessible and rich in antigen presenting cells, making it a preferred target site for immunization. To enable Ad-based clinical immunization, we developed microneedle arrays (MNAs) to deliver live Ad-vectors to the skin. The MNA design and fabrication process we developed results in MNAs that effectively preserve Ad viability, and enable efficient delivery of infectious vector to the skin. Specifically, MNA embedded live Ad5 virus retained infectivity for up to a year. Topical application of MNAs resulted in transduction of skin cells. Further, immunization of mice with an Ad5 expression vector encoding the SIVmac239 gag antigen resulted in the generation of potent CTL activity against the dominant gag 76-84 eptiope. Taken together, our results demonstrate the capacity of MNAs to preserve and deliver Ad5 virus to the skin for genetic immunization. This novel technology platform may enable a wide range of topical skin-targeted Ad vaccines.