Transmission blocking vaccines are one of the control strategies for vector-transmitted protozoan diseases. Antibodies raised in the vaccinated host prevent the development of the parasite in the insect vector, interrupting the epidemiological cycle. The FML antigen of Leishmania donovani in combination with saponin (FML-vaccine and Leishmune) induced 92-97% of protections against zoonotic visceral leishmaniasis. We assayed the ability of FML to inhibit Leishmania donovani and Leishmania chagasi procyclic promastigote-binding to dissected Lutzomyia longipalpis midguts. We found a dose-dependent inhibition, more pronounced on L. donovani (80%) than on L. chagasi promastigotes (p<0.001). On the other hand, the Fab-IgG serum fraction of Leishmune vaccinated dogs (IgG2 predominant), also inhibited parasite binding in a dose-response (p<0.0001) with an equally potent effect against L. donovani or L. chagasi (p = 0.061). The transmission blocking properties of the Leishmune vaccine was also assessed by an in vivo membrane assay, with sand flies fed with 1.5 x 10(7) amastigotes, human blood and, vaccinated or normal control dog sera. Significantly higher values were found in rate of infection (p<0.025) and intensity of infection (number of parasites/insect) (p<0.05) of control sand flies, making a very reduced infection index (20.7%) in the vaccine group. Our results disclosed that the Leishmune vaccine is a TBV, and that the dog antibodies present in sera, even 12 months after vaccination, lead to a significant effective protection of 79.3%.