The performance of PCR to detect Leishmania kDNA in serum for the diagnosis of visceral leishmaniasis (VL) was assessed in serum samples from 65 patients with VL, 17 non-infected individuals and 17 patients with other febrile hepatosplenic diseases. Serum PCR showed a sensitivity of 85%, specificity of 100% and efficiency of 90%. The sensitivity values obtained for blood PCR (97%) and rK39 ELISA (95%) were significantly higher (P=0.01) than the values observed for L. chagasi ELISA (88%) and serum PCR (85%), whilst no difference was observed among the specificity rates obtained with rK39 ELISA (94%; P=0.47) and L. chagasi ELISA (85%; P=0.06). This work suggests that the use of serum samples may be an alternative for the diagnosis of VL when peripheral blood samples are not available or require significant operational efforts. (C) 2008 Published by Elsevier Ltd on behalf of Royal Society of Tropical Medicine and Hygiene