We have reviewed our experience with 271 B-cell lymphomas to determine the effectiveness of flow cytometry in the characterization of these malignancies. Flow cytometric immunophenotyping of the total lymphocyte and/or large lymphocyte populations confirmed the morphologic diagnosis of B-cell lymphoma in 92% of cases, which included 79% monoclonal and 13% surface immunoglobulin (SIg)-negative lymphomas. Light chain monoclonality was most frequent in low grade and follicular center cell (FCC) lymphomas, while SIg-negative cases were most common in high grade and non-FCC types. Low grade lymphomas of all histologic types had high median percentages of neoplastic cells and low T-cell percentages. Conversely, high grade lymphomas exhibited lower and less uniform median percentages of B-cells, with higher numbers of T-cells and lower CD4/CD8 ratios than low grade lymphomas. Several differences in B- and T-cells were observed between specific high grade histologic types. FCC lymphomas with a diffuse pattern had lower percentages of CD4+ cells and lower CD4/CD8 ratios than cases with a follicular pattern. Thus, immunophenotypic differences were observed between histologic types or groups with known differences in clinical course and prognosis. We conclude that flow cytometry provides reliable information on neoplastic and nonneoplastic cells in lymph nodes involved by B-cell lymphomas.