A total of 135 fine-needle aspiration (FNA) biopsies from varying sites were performed in 123 children (mean, 10.5 years; range, one day to 18 years) over a five-year period. One hundred thirty (96.3%) biopsy specimens were satisfactory for evaluation. Seventy-nine cases were nonneoplastic (60.8%); among these cases, a specific diagnosis of infectious disease was made in 17 (13.1%). A diagnosis of neoplastic disease was made in 50 (38.5%) cases, of which 14 (10.8%) were benign, 28 (21.5%) were malignant, and 8 (6.2%) were neoplasms of uncertain biologic potential. The sensitivity of pediatric FNA biopsies was 90.6%, specificity 100%, positive predictive value 100%, negative predictive value 94.7%, and efficiency of the test 96.5%. There were no false-positive diagnoses and there were four false-negative diagnoses, three of which involved aspirates of the central nervous system (CNS). Ancillary studies, including immunocytochemistry (20 cases), electron microscopic examination (18 cases), microbiologic culture (8 cases), cytogenetic studies (7 cases), and flow cytometry (3 cases), were performed on the aspirated material, enabling a more specific diagnosis or supplying additional information in many cases. Definitive diagnosis by FNA biopsy enabled radiation therapy and/or chemotherapy to be administered for unresectable malignant neoplasms, provided material for culture of infectious lesions, identified benign lesions not needing surgery, and aided the surgeon in planning the extent of surgery in resectable malignant neoplasms. These results support the greater use of FNA biopsy in the pediatric population.