DL1 generates a microenvironment that is progenitor cell expansion supportive. (A) The addition of DL1 to the fed-batch culture system enhanced the expansion of (i) CD34⁺ cells; (ii) CD34⁺CD90⁺ cells; (iii) CFCs; (iv) LTC-ICs. (Bi) Addition of DL1 to HSC-enriched (Lin⁻Rho^loCD34⁺CD38⁻CD45RA⁻CD49f⁺) cells did not lead to a significant increase in the expansion of CD34⁺CD90⁺ cells after 7 days of culture. (Bii) In contrast, the addition of DL1 to CD34⁺ cells for 16 days significantly enhanced CD34⁺CD90⁺ expansion as compared with cells cultured without DL1. (C) Microarray analysis indicated that the Notch1 and Notch2 receptors are expressed at varying levels in many hematopoietic populations. (D) DL1 caused lineage skewing in hematopoietic cell culture, most notably a decrease in CD14⁺ cells and CD15⁺ cells, and an increase in CD7⁺ cells. (E) When CD14⁺ cells or their CM were cultured with Lin⁻Rho^loCD34⁺CD38⁻CD45RA⁻CD49f⁺ cells, there was an inhibitory effect on the proliferation of the CD34⁺CD90⁺ population. (F) The CM from the specified mature cell lineages (CD14⁺, CD15⁺, CD41⁺, GlyA⁺, CD7⁺, other = CD34⁻CD14⁻CD15⁻CD41⁻GyA⁻CD7⁻) was added to the Lin⁻Rho^loCD34⁺CD38⁻CD45RA⁻CD49f⁺ cells and cultured for 7 days. Different lineages had varying impacts on the expansion of (i) total cells and (ii) CD34⁺CD90⁺ cells. Effects were categorized as negative (red region), neutral (gray region), or positive (green region) as compared with CM from unsorted cell populations (all). All error bars indicate standard deviation. n ≥ 3, *P < .05. CFU-E, CFU-erythroid; CFU-M, CFU-macrophage; CFU-G, CFU-granulocyte; CFU-MK, CFU-megakaryocyte; CMP, common myeloid progenitor; ERY, erythrocyte; GMP, granulocyte/monocyte progenitor; MEGA, megakaryocyte; MEP, megakaryocyte/erythrocyte progenitor; MLP, multilymphoid progenitor; MONO, monocyte; NEUT, neutrophil.