Some B-cell lymphoproliferative disorders displaying a serum monoclonal immunoglobulin (Ig) M protein could be difficult to differentiate from Waldenström's macroglobulinemia (WM). We report on the immunophenotypic and cytogenetic characteristics of 85 patients with WM and compare them with 29 patients with splenic marginal zone lymphoma (SMZL). For immunophenotyping, WM and SMZL constantly expressed pan–B-cell markers (CD19, CD20, CD22, and surface Ig). However, there were differences in the κ/λ ratio (1.2:1 for SMZL and 4.5:1 for WM) and in some markers such as CD22 and CD11c, which were overexpressed in patients with SMZL compared with patients with WM, whereas CD25 was more frequently positive in WM (88% vs. 44%). The CD103 antigen was always negative in WM, whereas it was positive in 40% of SMZL cases. The monoclonal antibody FMC7 was usually positive in both entities: heterogeneous in WM but homogeneous in SMZL. The combination of CD25 and CD22 could differentiate between WM and SMZL. The principal molecular abnormality in WM is 6q deletion (30% in our experience), whereas in SMZL the most common abnormalities are loss of 7q (19%) along with +3q (19%) and +5q (10%). Interestingly, the incidence of IgH rearrangement was low in WM (12%) and SMZL (10%). Immunophenotypic and molecular cytogenetic studies could help to distinguish WM from SMZL.