Biliary atresia is the commonest cause of cholestatic jaundice. Early diagnosis & surgical correction helps long time survival & prevent development of cirrhosis. This study was conducted to find out the role of liver biopsy in the diagnosis of biliary atresia having positive hepatobiliary scintigraphy as there is chance of false positivity. This cross sectional study was carried out in Bangabandhu Sheikh Mujib Medical University (BSMMU), Dhaka, Bangladesh at Paediatric Gastroenterology & Nutrition Department from January 2014 to June 2015. All admitted patients with the diagnosis of neonatal cholestasis were evaluated clinically and by scintigraphy for biliary atresia. Having positive hepatobiliary scintigraphy but highly suggestive of biliary atresia were enrolled for this study. A total of 108 cases were initially selected. Among them 33 patients showed no excretion of tracer during hepatobiliary scintigraphy. Liver biopsy was done in these 33 cases. All cases (100%) had history of passage of pale or acholic stool. Fractionated serum bilirubin of >2mg/dl was found all of the cases. Histological scoring system reveals typical features of biliary atresia in 27(81.8%). Two had no conclusive histology of biliary atresia, four had features of neonatal hepatitis (12.1%). In this study, percutaneous liver biopsy with histological analysis by scoring system was found useful for the correct diagnosis of biliary atresia.