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Hepatobiliary scintigraphy (HBS) is an important investigation for the diagnosis of biliary atresia (BA) and its differentiation from causes of conjugated hyperbilirubinemia that do not require surgical intervention. Delayed imaging at 24 hours and phenobarbitone augmentation for 5 days has been required to achieve high sensitivity and specificity with current techniques. This study explores whether adding single photon emission computed tomography (SPECT) performs as well as existing methods without requiring delayed 24-hour imaging and whether the phenobarbitone premedication is necessary in all cases. Methods: A retrospective analysis of 105 HBS studies on 94 patients was performed. HBS included SPECT at 4 to 6 hours postinjection when no tracer was seen in the gastrointestinal tract in the first 60 minutes. This was done in 80 patients. Results: Gastrointestinal activity was seen in 14 patients within 60 minutes. For 4- to 6-hour studies, standard HBS and HBS with SPECT data showed a sensitivity of 100% for the diagnosis of BA. The specificity, accuracy, and positive likelihood ratios (PLR) were 67%, 75%, and 3 (confidence interval [CI] = 2.03-4.16) for planar imaging at 4 to 6 hours and 90%, 93%, and 10 (CI = 4.42-19) for 4- to 6-hour planar and SPECT imaging. When the 11 patients who had phenobarbitone stimulation were included, the results improved to 97%, 98%, and 30 (CI = 7.06-80). Conclusion: The addition of SPECT 4 to 6 hours postinjection of tracer significantly improves the diagnostic accuracy of hepatobiliary scintigraphy compared with planar imaging alone. This accuracy is as good as HBS performed after phenobarbitone stimulation. The combined technique of HBS with SPECT and phenobarbitone has the highest accuracy. Delayed imaging at 24 hours is usually not necessary.