CBA/N mice and F1 male mice, which are hemizygous for the CBA/N X chromosome, have an immune defect which is associated with the absence (deficiency) of a subpopulation of mature or late developing B lymphocytes. This characteristic was utilized to develop an antiserum that was specific for this subclass of B cells. C57BL/L mice were immunized with DBA/2 spleen calls, and the resulting antisera was absorbed with lymphoid cells derived from immunologically abnormal (CBA/N female X DBA/2 male)F1 male mice. The absorbed antisera was cytotoxic for a subpopulation of lymphocytes that was present in the spleens of adult DBA/2 and (CBA/N female X DBA/2 male)F1 female mice. The cells killed by the absorbed antisera were Ig-bearing, complement receptor-bearing B lymphocytes, which had a low-to-intermediate density of total surface Ig. Moreover, the cells remaining after treatment of adult (CBA/N female X DBA/2 male)F1 female spleen cells with the absorbed antisera and C had a high ratio of surface IgM to IgD. The development of this cytotoxic alloantisera, which is specific for a late developing (mature) subpopulation of B lymphocytes, will allow the functional characterization of subclasses of B lymphocytes.