Determination of successful vaccination with vaccinia virus is based on visual confirmation of a dermal response (take). Some revaccinees do not manifest a take, which may be due to a preexisting immunity rather than to poor technique or inadequate virus. Cytotoxic T-lymphocyte (CTL) response appears to be the most important immune defense in limiting response to vaccination. We evaluated vaccinia virus-specific CTL responses in revaccinees. Subjects with and without takes displayed comparable CTL responses. Vaccinia virus-specific CD8+ CTL responses may be useful in interpreting the response to vaccination, particularly in individuals who are revaccinated and have difficult-to-interpret visual takes.