This modified lectin affinity electrophoresis method is suitable for simultaneous measurement of liver, bone, and high-molecular-mass (high-Mr) isoforms of alkaline phosphatase (ALP; EC 3.1.3.1) in children. Age-related isoform reference ranges were derived for 247 children, ages 0-13 years. Liver ALP did not appear in plasma until after six months of age, and remained constant after one year of age. Bone ALP was highest in the youngest age group, and declined to relatively constant activities thereafter. High-MrALP was not detected in normal children. In bone disease, the bone isoform was increased in all age groups. In liver disease, only 5 of 30 infants younger than six months had detectable liver ALP, although half had increased bone ALP. Among children older than six months, 5 patients with biliary atresia and 15 patients with hepatitis A all had increased liver isoform activity. Liver ALP was also a sensitive index of early hepatobiliary complications in 27 nonjaundiced children with cystic fibrosis. Measurement of ALP isoforms therefore yields useful clinical information in children older than six months but is of doubtful value in younger infants.