The long-term bone marrow culture system developed by Dexter (MBMC) is known to store immature lymphoid precursors capable of differentiating into mature B cells in irradiated or immunodeficient mice. It has been suggested that pre-B cells are not generated under such culture conditions, but that opinion was not based on any systematic analyses. In the present study under carefully controlled conditions, we observed that pre-B and pro-B cells were eliminated from the late stage of primary MBMC, and the former were not generated in recharged MBMC. Under appropriate conditions, these immature precursors in recharged MBMC generated in vitro immunoglobulin-positive (Ig+) cells to differentiate into antibody-forming cells upon stimulation with lipopolysaccharide (LPS). LPS-reactive B cells were observed in every 10th of the Ig+ cells, the frequency being essentially the same as that observed in normal B cells in different tissues. The immature B cell precursors generating LPS reactive cells were expressed in recharged MBMC at the frequency of 4.2 x 10(-6). A staining experiment showed that cells bearing AA4.1 were stored at the frequency of 10(-4)-10(-5). This frequency is thought to be similar to that of lymphoid precursors in recharged MBMC committed to differentiate along B lineage cells. Based on these results, we discussed the stage, nature, and mode of differentiation of immature lymphoid precursors stored in MBMC.