Abstract: Dot-ELISA for monoclonal antibodies specific to different species of leishmania promastigotes from naturally infected sandflies was described. One of the monoclonal antibodies, L9E12 was selected for further analysis. The highest titer of the ascites fluid With promastigotes of Xinjiang strain determined by the Dot-ELISA was 1:655 360. The promastigotes Zi 10 and Zi 11 tested with hybridoma L9E4 ascites fluid showed a titer of 1:81920 dilution which was 3-20 times higher than by the IFA technic.So the Dot-ELISA described is proved to be very sensitive, specific and fast in the determination of promastigotes species from naturally infected sandflies.