Tracking hematopoiesis at the single cell level.

Despite intensive research, many longstanding questions of experimental hematology remain unsolved. One major reason is the fact that hematopoiesis is usually followed by analyzing populations of cells rather than individual cells, at few points in time during an experiment and without knowing (or quickly loosing) the cells' individual identities. The static picture yielded by this approach makes it impossible to appreciate the dynamic developmental processes leading to the generation of the full hematopoietic system from individual hematopoietic stem cells (HSCs). Real-time tracking of individual cells in culture, tissues, or whole organisms would be an extremely powerful approach to fully understand the developmental complexity of hematopoiesis. To this end, a computer-aided culture and bioimaging system is being developed to follow the fate of individual cells over long periods of time. This system is used to follow the development of multilineage cobblestone colonies from adult HSCs in stroma cocultures at the single cell level over many generations. To facilitate noninvasive detection of lineage commitment in these cultures, new subcellular forms of optimized fluorescent proteins have been developed to allow simultaneous marking of multiple hematopoietic lineages within the same animal.