Chronic lymphocytic leukemia (CLL) lymphocytes manifest anomalous motility and cap formation. Since these processes involve cytoskeletal proteins, vimentin from intermediate filaments of normal and CLL lymphocytes was investigated using hetero- and monoclonal antisera. The antisera reacted predominantly with a 60-kD polypeptide, following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of total lymphocyte proteins. When lymphocytes were stained by indirect immunofluorescence, normal lymphocytes demonstrated well defined cytoplasmic fibrils that capped spontaneously after contact with a glass surface and incubation at 37 degrees C. This capping was dependent on energy and intact microfilaments. Lymphocytes from patients with CLL showed several patterns. In one group, the initial staining was weak, and few capped cells were present after incubation. Lymphocytes from other patients had either normal or aberrantly organized fibrils in which capping was diminished. In another group, a fibrillar pattern with normal or increased capping was seen. In total, 47% +/- 5.1% (mean +/- SE) of normal lymphocytes capped after a 1-hr incubation at 37 degrees C (n = 12) compared to 21% +/- 5.1% for CLL lymphocytes (n = 20, p less than 0.002). Purified subpopulations of normal B and T cells did not differ from unfractionated normal lymphocyte populations. These results demonstrate an anomalous vimentin capping response in CLL lymphocytes. They also show that the arrangement of vimentin in these cells differs from that of normal lymphocytes.