The neural cell adhesion molecule L1 is a cell surface glycoprotein of the immunoglobulin superfamily which mediates adhesion between neural cells. The possibility that similar cell-cell recognition mechanisms may be shared by the nervous and immune systems prompted us to study the expression and function of L1 in cells of the hematopoietic system. Immunofluorescence analysis using monoclonal L1 antibody revealed that the molecule is expressed in the bone marrow, spleen, and thymus of the mouse. This observation was confirmed by amplifying cDNA derived from these organs by the polymerase chain reaction with L1-specific oligonucleotide primers. Two-color fluorescence analysis indicated that bone marrow lymphoid and granulocyte precursor cells express low and high levels of L1, respectively. In the thymus L1 is primarily expressed by mature cells that have a strong expression of CD3 and in the spleen both B cells and T cells express L1. The possible function of L1 in lymphoid cells was studied using subcloned ESb-MP lymphoma cells having high or low densities of L1 on the cell surface as well as activated splenic B lymphoblasts. Parental and subcloned ESb-MP cells that strongly expressed L1 could form homotypic aggregates in the presence of low Ca2+ levels, whereas subcloned ESb-MP cells with a weak expression of L1 did not aggregate, suggesting that L1 mediates the Ca(2+)-independent aggregation of the parental ESb-MP cells. Furthermore, the aggregation of activated B lymphoblasts under physiological concentrations of Ca2+ and Mg2+ was inhibited by 30% in the presence of Fab fragments of polyclonal L1 antibodies, implying that L1 also mediates adhesion among normal lymphoid cells. A possible role of L1 on lymphocytes in stimulating the innervation of lymphoid organs is discussed.