Acute lymphoblastic leukemia (ALL) is the most frequent cancer encountered in children. Little is known about the molecular basis of childhood ALL, although the clinical, pathological, and immunophenotypic features have been well documented. To understand the role of tumor suppressor genes (TSGs) in the development of this disease, we performed a detailed allelotype analysis. Twenty-nine patients (24 pre-B and 5 of T-cell lineage) were investigated for loss of heterozygosity (LOH), using 49 highly polymorphic markers distributed over 13 chromosomal arms which are known or postulated to contain TSGs. The highest rates of allelic losses were observed in chromosomes 9p and 12p which were deleted in 29 and 32% of the informative patients, respectively. These are among the most frequent alterations found in childhood ALL. Other losses were found at a lower frequency in chromosomes 6p, 6q, 9q, 17p, and 17q. No LOH was found at chromosomes 3p, 5q, 11p, 11q, 13q and 18q in any patient. These results suggest that many TSGs may be involved in the development of childhood ALL.