Cells in colonies of culture-derived mouse and rat bone marrow macrophages were examined for membrane Ia antigen to determine if the steady-state expression of the antigen was restricted to macrophages derived from a distinct population of progenitor cells. Multiple subcultures of macrophages derived from single soft-agar colonies were tested for Ia+ cells on three different days of culture. About one-half of the colonies gave rise to subcultures that never contained Ia+ cells, about 40% yielded subcultures that contained some Ia+ cells on at least 2 of the 3 assay days, and about 10% of the colonies produced subcultures that contained Ia-bearing cells on all 3 assay days. Thus, when cultures were assayed at any one time for their content of Ia-bearing cells, the results raised the possibility of phenotypically distinct subpopulations of progenitors. However, sequential analyses of the subcultures revealed the variable expression of Ia on cells from at least one-half of the colonies. We conclude that, under steady-state conditions, the presence of Ia antigen on bone marrow-derived macrophages is not clonally restricted. That a T-cell-derived lymphokine induced Ia antigen on essentially all the cells in most of the colonies of macrophages confirms their potential to express the antigen.