Six new B lineage lymphomas of NFS mice established in primary tissue culture were examined for a number of phenotypic, functional, virologic, and molecular genetic characteristics. Two of the tumors and their cloned derivatives bore surface markers characteristic of B cells, whereas four tumors resembled pre-B cells. One of the B cell and two of the pre-B cell lymphomas had rearrangements of both heavy and light chain immunoglobulin genes, confirming their designation as B-lineage lymphomas. All the tumors but one were Ly-1+, indicating that Ly-1 may be expressed by some pre-B cells as well as some B cells. In addition, one pre-B cell lymphoma was Mac-1+. MCF murine leukemia viruses obtained from two of the tumors did not accelerate development of B-lineage lymphomas in NFS mice.