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Although endoscopy with biopsy is the gold standard for the diagnosis of inflammatory bowel disease (IBD), this procedure is invasive and its repetition is stressful, especially in children. The purpose of this study was to evaluate prospectively the role of (99m)Tc-HMPAO-labeled granulocyte scintigraphy in the diagnosis and follow-up of pediatric IBD and its possible use as an alternative to colonoscopy to determine the disease extent and severity beyond its ability to differentiate ulcerative colitis from Crohn's disease. During a 10-year period, 52 children, aging between 2 and 17 years (median, 11.09 years), were subjected to (99m)Tc-HMPAO granulocyte scan 7-28 days from conventional diagnostic tests, and the results were compared with endoscopic and bioptic results. Disease severity was graded by the focal uptake intensity versus iliac bone uptake (Scan Activity Index) and compared with Endoscopy Mayo Score. In 15 out of 16 patients, IBD diagnosis was obtained with a full correspondence of location and severity of lesions, respectively, in 14 out of 16 and 13 out of 16, while in 31 out of 36 patients, IBD was correctly excluded (sensitivity of 93.7%, specificity of 86.1%, and negative predictive value of 96.4%). During the follow-up, all relapses (24) and remissions (13) were correctly recognized (sensibility and specificity of 100%). In conclusion, (99m)Tc-HMPAO granulocyte scan is an accurate minimally invasive technique with very good accuracy, able to diagnose and to grade the intensity and extent of the disease; it is also a useful tool in the follow-up of pediatric IBD where it could reliably replace the invasive endoscopic assessment in most cases.