Infection with reovirus 3 (Reo‐3) has been suggested as the cause of extrahepatic biliary atresia and idiopathic neonatal hepatitis but confirmation has been lacking. Therefore we have searched for a specific anti‐Reo‐3 antibody response in the sera of patients with biliary atresia or neonatal hepatitis and for Reo‐3 antigens in their hepatobiliary tissues. Sera from 23 infants with extrahepatic biliary atresia, 12 with neonatal hepatitis, 30 age‐matched control patients with other liver diseases, and 55 control patients without liver disease were tested by an enzyme‐linked immunosorbent assay for total (IgA, IgG, and IgM) anti‐Reo‐antibodies; sera of infants younger than 6 months of age were tested also for IgM anti‐Reo‐3 antibodies alone. There was no difference between either total or IgM anti‐Reo‐3 antibody levels in infants with extrahepatic biliary atresia or neonatal hepatitis and levels in control infants. Reo‐3 antigens were not detected in the hepatobiliary tissues of 19 infants (18 with biliary atresia, one with neonatal hepatitis) by an immunoperoxidase method that readily demonstrated Reo‐3 in control in fected HEp‐G2 cells. Our data do not support a relationship between neonatal liver diseases and infection with Reo‐3.