This prospective study was done to assess the frequency of acute pyelonephritis (APN) in febrile children with positive urine culture as documented by Tc99m DMSA scintigraphy (DMSA) and the frequency of vesicoureteric reflux (VUR) in these children. Secondly, to determine the frequency of APN, in febrile children with supportive evidence for UTI but with negative urine culture, as documented by DMSA and frequency of VUR in them. Thirdly to stress the utility of DMSA to diagnose APN in urine culture negative febrile children and to suggest DMSA as a clinical tool in evaluation of fever of unknown origin (FUO). This study included 42 children with positive urine culture and 26 children with negative urine culture who had supportive evidence of UTI as determined by the predetermined criteria and diagnosed to have APN by DMSA. All of them had ultrasonogram (USG), DMSA and voiding cystourethrogram (VCU). They were followed up for a minimum period of 6 months. Out of the 42 children with positive urine culture 92.9% had features of APN in the DMSA of whom 82.1% had vesicoureteric relux (VUR). The DMSA was abnormal in 26 children with negative urine culture, of whom 65.4% had VUR. Ultrasound suggestive of parenchymal change was observed in 47.6% in the culture positive group and 65.4% in the culture negative group. In conclusion, it is suggested, that DMSA is a useful investigation for the diagnosis of APN in febrile UTI. DMSA is indicated in febrile children with negative urine culture but with supportive evidence of UTI and in FUO. An abnormal DMSA is a strong indication for work up for VUR.