Introduction
Hematopoietic stem cells (HSC) are well-characterized, tissue-specific stem cells that exhibit
remarkable self-renewal capacity and are responsible for the life-long maintenance of the
hematopoietic system. HSC are rare cells that reside in adult bone marrow where hematopoiesis is
continuously taking place, but can also be found in cord blood, fetal liver, adult spleen and peripheral
blood. Throughout the life span, hematopoiesis continuously replenishes the various lymphoid,
myeloid and erythroid-megakaryocyte lineages, but also to maintain a small pool of HSC with the selfrenewal
capacity that is capable of carrying on hematopoiesis. From HSC to mature blood cells,
extensive proliferation and expansion occurs that results in the production of millions of blood cells.
When cultured in a suitable semi-solid matrix, such as methylcellulose supplemented with nutrients
and cytokines, HSC or hematopoietic progenitors called colony-forming cells (CFCs) proliferate to
form discrete cell clusters or colonies. Colony types include colony-forming unit-erythroid (CFU-E),
burst-forming unit-erythroid (BFU-E), CFU-granulocyte, macrophage (CFU-GM) and CFUgranulocyte,
erythrocyte, macrophage, megakaryocyte (CFU-GEMM).
In classical CFC assays, culture usually takes place in a 35 mm dish for 14-21 days for the colonies to
reach certain size (> 40 cells/colony) for manual counting. The CFCs are classified and manually
enumerated based on the morphological recognition of one or more types of hematopoietic lineage
cells within the colony.
Cell Biolabs CytoSelect™ 96-well Hematopoietic Colony Forming Cell Assay does not involve
subjective manual counting of colonies or require a 2-3 week incubation period. Instead cells are
incubated only 7-10 days in a semisolid methycellulose media before being solubilized, lysed and
detected by the patented CyQuant® GR Dye in a fluorescence plate reader (see Assay Principle
below). Alternatively, viable CFCs can be easily recovered for further culturing and testing. This
format provides a quantitative, high-throughput method to accurately measure HSC or hematopoietic
progenitor clonogenic capability.