The aim of the study was to test a possible extension of the period between vaccination and revaccination with the live Trichoben vaccine (Bioveta, a.s., Czech Republic). The calves (n = 61) were vaccinated intramuscularly. The intervals between vaccination and re-vaccination were 3, 5, 7, 10, 14 or 28 days. Another group of calves (n = 16) was vaccinated once with a double dose of the vaccine. The protective immunity against experimental infection with a virulent Trichophyton verrucosum CCM F-650 strain was investigated in animals one month after the last vaccination and compared with a group of non-vaccinated controls (n = 16). Clinical results were evaluated and compared using balanced two-way-ANOVA. The level of post-vaccination immunity in cattle after re-vaccination was sufficient when the intervals were 5, 7, 10, 14 and 28 days. One-time injection of a double prophylactic dose of the vaccine and a 3-day interval between vaccinations reduced the immunity and vaccine efficacy to an unsatisfactory degree. There were significant differences between the groups of calves vaccinated using an interval longer than 5 days, and the group vaccinated once with a double dose, and the group with a 3-day interval between vaccinations. The level of post-vaccination immunity in cattle after re-vaccination at the commonly used interval of 10 -14 days was identical with the immunity achieved when the interval was reduced to 5 or 7 days or extended up to 28 days. The extension of the interval between vaccinations to 5 -28 days is recommended for using the Trichoben vaccine in veterinary practice.