Hematopoietic stem and progenitor mobilization has revolutionized the field of hematopoietic transplantation. Currently, hematopoietic grafts acquired from the peripheral blood of patients or donors treated with granulocyte-colony stimulating factor (G-CSF) are the preferred source for transplantation. G-CSF mobilization regimens, however, are associated with known morbidities and a significant number of normal donors and patient populations fail to mobilize sufficient numbers of hematopoietic stem and progenitor cells for transplantation, necessitating the need for non-G-CSF mobilization strategies. Mechanistic studies evaluating hematopoietic bone marrow niche interactions have uncovered novel agents with the capacity for hematopoietic mobilization. This chapter provides a comprehensive overview of mobilizing agents, other than G-CSF, and experimental procedures and technical aspects important to evaluate and define their hematopoietic mobilizing activities alone and in combination.