ABSTRACT Previous studies have demonstrated that protection against New World leishmaniasis caused by Leishmania amazonensis can be elicited by immunization with the developmentally regulated Leishmania amastigote antigen, P-8. In this study, several independent experimental approaches were employed to investigate the protective immunological mechanisms involved. T-cell subset depletion experiments clearly indicate that elicitation of CD8 ⁺ (as well as CD4 ⁺ ) effector responses is required for protection. Further, mice lacking β ₂ -microglobulin (and hence deficient in major histocompatibility complex class I antigen presentation) were not able to control a challenge infection after vaccination, indicating an essential protective role for CD8 ⁺ T effector responses. Analysis of the events ongoing at the cutaneous site of infection indicated a changing cellular dynamic involved in protection. Early postinfection in protectively vaccinated mice, a predominance of CD8 ⁺ T cells, secreting gamma interferon (IFN-γ) and expressing perforin, was observed at the site of infection; subsequently, activated CD4 ⁺ T cells producing IFN-γ were primarily found. As protection correlated with the ratio of total IFN-γ-producing cells (CD4 ⁺ and CD8 ⁺ T cells) to macrophages found at the site of infection, a role for IFN-γ was evident; in addition, vaccination of IFN-γ-deficient mice failed to provide protection. To further assess the effector mechanisms that mediate protection, mice deficient in perforin synthesis were examined. Perforin-deficient mice vaccinated with the P-8 antigen were unable to control infection. Thus, the elicitation of CD8 ⁺ T cell effector mechanisms (perforin, IFN-γ) are clearly required in the protective immune response against L. amazonensis infection in vaccinated mice.