Recent advances in our understanding of the earliest stages of hematopoietic cell differentiation, and how these may be manipulated under defined conditions in vitro, have set the stage for the development of robust bioprocess technology applicable to hematopoietic cells. Sensitive and specific assays now exist for measuring the frequency of hematopoietic stem cells with long-term in vivo repopulating activity from human as well as murine sources. The production of natural or engineered ligands through recombinant DNA and/or combinatorial chemistry strategies is providing new reagents for enhancing the productivity of hematopoietic cell cultures. Multifactorial and dose-response analyses have yielded new insight into the different types and concentrations of factors required to optimize the rate and the extent of amplification of specific subpopulations of primitive hematopoietic cells. In addition, the rate of cytokine depletion from the medium has also been found to be dependent on the types of cell present. The discovery of these cell-type-specific parameters affecting cytokine concentrations and responses has introduced a new level of complexity into the design of optimized hematopoietic bioprocess systems.