Cells derived from lymph nodes (LN) of adult CBA mice respond blastogenically to mitomycin-treated autologous, as well as isologous spleen cells. This isogeneic LN-to-spleen (mixed lymphocyte culture) is best obtained when both responder and stimulator cells are derived from donors greater than 10 weeks of age. Responsive cells appear restricted to LN since they could not be detected in adult spleen, marrow, or thymus. LN cells do not require the presence of spleen in order to differentiate into responder cells since those derived from neonatally splenectomized mice are fully active. Stimulator cells appear in the spleen, bear Ig on their surfaces, and can be detected in spleens of irradiated, bone marrow-reconstituted mice. Experiments comparing the responsiveness of adult LN cells and that of neonatal T cells toward mitomycin C-treated lymphoid cells from a variety of sources suggest the presence of two iso-antigens on B lymphocytes. Since both antigens apparently are absent on precursor bone marrow cells and develop with time, they have been classified as murine differentiation antigens 1 and 2 (MDA-1, MDA-2). Whereas both appear in the spleen, only one, MDA-1, is also detectable by this methodology in LN. Both MDA-1 and MDA-2 activate neonatal T cells, but MDA-2 triggers only adult LN. Whereas MDA-2 developed in an x-irradiated, bone marrow-reconstituted spleen, MDA-1 did not over a 9-week interval.