We assessed an inactivated whole virus feline leukemia virus (FeLV) vaccine developed from a molecularly cloned feline leukemia virus isolate (FeLV-61E-A) for its ability to protect cats against homologous and heterologous virulent virus challenge. The fractions of cats that resisted the induction of persistent viremia after FeLV challenge were the following: (1) FeLV-61E-A vaccine, 95%; (2) adjuvant controls, 26%; and (3) established commercial control FeLV vaccine, 35%. The pre-challenge mean neutralizing antibody titers for each group were (1) FeLV-61E-A vaccine, 1:43; (2) adjuvant controls, <1:8; and (3) established commercial control FeLV vaccine: 1:12. The commercial version of the prototype FeLV-61E-A vaccine (Fel-O-Vax, Fort Dodge Laboratories, Fort Dodge, IA) was developed through use of a proprietary adjuvant and a stable high antigen production cell lines. The efficacy and duration of immunity produced by Fel-O-Vax was studied alone and in multivalent combination with other feline virus vaccines in seven subsequent efficacy trials conducted in over 150 immunized cats. The overall FeLV-resistant fraction in these trials was 87% in vaccinated cats versus 8% in adjuvant controls. The duration of protective immunity induced by the multivalent Fel-O-Vax-LvK IV at 1 year postvaccination was 100% in challenged vaccinees versus 0% in challenged controls. The results of these studies show that an inactivated FeLV vaccine prepared from a molecularly cloned subgroup A FeLV can produce high level protective immunity against FeLV infection. This immunity is durable for at least 1 year without intervening booster immunization or virus exposure.