Allelic loss at nonrandom chromosomal sites is thought to mark the position of tumor suppressor genes involved in the pathogenesis and progression of human malignancies. Solid tumors in particular have been found to harbor multiple genetic changes resulting in loss of function mutations. Tumor suppressor genes have also been found to be involved in the progression of lymphoid tumors. Previous reports have suggested the involvement of a tumor suppressor gene located on the long arm of chromosome 13, between the retinoblastoma (RB) and D13S25 loci, in the pathogenesis and or progression of more than 40% of B-cell chronic lymphocytic leukemia (B-CLL), a common lymphoid malignancy whose molecular etiology remains largely unknown. In the present study, we report the construction and characterization of a YAC contig spanning a region of approximately 3 cM between the RB gene and the D13S31 locus. We also screened 60 paired normal/tumor B-CLL samples for allelic loss on chromosome 13 with nine microsatellite markers located between RB and D13S25. This analysis has allowed us to narrow the smallest region of loss to a segment of 550 kb located between the 206XF12 and D13S25 markers.