Cytotoxic T lymphocytes (CTL) play a vital role in host defense against viral and intracellular bacterial infections. However, nonreplicating vaccines administered by intramuscular injection using a syringe and needle elicit predominantly humoral responses and not CTL responses. Here we report that epidermal powder immunization (EPI), a technology that delivers antigens on 1.5- to 2.5-μm gold particles to the epidermis using a needle-free powder delivery system, elicits CTL responses to nonreplicating antigens. Following EPI, a majority of the antigen-coated gold particles were found in the viable epidermis in the histological sections of the target skin. Further studies using transmission electron microscopy revealed the intracellular localization of the gold particles. Many Langerhans cells (LCs) at the vaccination site contained antigen-coated particles, as revealed by two-color immunofluorescence microscopy, and these cells were found in the draining lymph nodes 20 h later. Immune responses to several viral protein antigens after EPI were studied in mice. EPI with hepatitis B surface antigen (HBsAg) and a synthetic peptide of influenza virus nucleoprotein (NP peptide) elicited antigen-specific CTL responses as well as antibody responses. In an in vitro cell depletion experiment, we demonstrated that the CTL activity against HBsAg elicited by EPI was attributed to CD8⁺, not CD4⁺, T cells. As controls, needle injections of HBsAg or the NP peptide into deeper tissues elicited solely antibody, not CTL, responses. We further demonstrated that EPI with inactivated A/Aichi/68 (H3N2) or A/Sydney/97 (H3N2) influenza virus elicited complete protection against a mouse-adapted A/Aichi/68 virus. In summary, EPI directly delivers protein antigens to the cytosol of the LCs in the skin and elicits both cellular and antibody responses.