STEMdiff™ Hematopoietic Kit
For differentiation of human ES or iPS cells into hematopoietic progenitor cells
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What Our Scientist Says
We developed the STEMdiff™ Hematopoietic Kit so scientists like you can have a reliable source of hPSC-derived hematopoietic progenitor cells for their research.
The robust, two-stage protocol initially induces cells toward the mesoderm before further differentiating them into hematopoietic progenitor cells. After 12 days, a population of hematopoietic cells containing 25 - 65% (average 43%) CD34+CD45+ progenitor cells can be harvested, including progenitor cells that have the capacity to form functional hematopoietic colonies in the colony-forming unit (CFU) assay.
STEMdiff™ Hematopoietic Kit has been optimized for differentiation of human pluripotent cells maintained in:
• mTeSR™1 (Catalog #85850)
• mTeSR™ Plus (Catalog #100-0276)
• TeSR™-E8™ (Catalog #05990)
Figure 1. Hematopoietic Differentiation Protocol
On Day -1, harvest and seed human ES/iPS cell colonies as small aggregates in mTeSR™1, mTeSR™ Plus, or TeSR™-E8. After one day, TeSR™ medium is replaced with Medium A (STEMdiff™ Hematopoietic Basal Medium containing Supplement A) to begin inducing the cells towards a mesoderm-like state (day 0). On day 2, a half medium change is performed with fresh Medium A. On day 3, the medium is changed to Medium B (STEMdiff™ Hematopoietic Basal Medium containing Supplement B) with half medium changes on days 5, 7 and 10, to promote further differentiation into hematopoietic cells. Typically, by day 12, large numbers of HPCs can be harvested from the culture supernatant.
Figure 2. Morphology of hPSC-Derived HPCs
Representative images of (A) hES (H1) cells and (B) hiPS (WLS-1C) cells on Day 12 of differentiation to HPCs using the STEMdiff™ Hematopoietic Kit. Differentiated cells exhibit typical HPC morphology as round cells that float freely in suspension.
Figure 3. Efficient and Robust Generation of CD34+CD45+/CD43+ HPCs
hES and hiPS cells were cultured for 12 days in single wells of 12-well plates using the STEMdiff™ Hematopoietic Kit. At the end of the culture period, cells in suspension were harvested and analyzed by flow cytometry for expression of hematopoietic cell surface markers: CD34, CD45 and CD43. (A,B) Example flow cytometry plots for hematopoietic cell surface-marker analysis of cultures of hES (H1 and H9) and hiPS (STiPS-M001) cells. (C,D) Percentages and total numbers of CD34+CD45+ cells in cultures of hES (H1 and H9) or hiPS (WLS-1C, STiPS-F016, STiPS-M001 and STiPS-B004) cells are shown. Data shown as mean ± SEM; n ≥ 3.
Figure 4. hPSC-Derived HPCs Produce Colonies of Multiple Lineages
Cells in suspension were harvested from the cultures on Day 12 of the hematopoietic differentiation protocol and assessed in colony-forming unit (CFU) assays using MethoCult™ H4435 Enriched (Catalog #04435) methylcellulose-based medium. (A) CFU frequencies in cultures of 6 different hPSC lines. (Data shown as mean ± SEM; n ≥ 3.) The CFU frequencies were variable between the different cell lines, with on average approximately 120 CFU per 10,000 hPSC-derived HPCs plated. (B) The progenitor cell types observed included granulocyte/macrophage (CFU-M, CFU-G and CFU-GM), erythroid (BFU-E and CFU-E) and occasional mixed (CFU-GEMM) colonies. Representative colony images are shown at 40X magnification.
Figure 5. Generation of Hematopoietic Progenitor Cells from hPSCs Maintained in mTeSR™ Plus
Human ES (H1, H9) and iPS (STiPS-M001, WLS-1C) cell lines maintained in mTeSR™1 (daily feeds) or mTeSR™ Plus (restricted feeds) were differentiated to hematopoietic progenitor cells using the STEMdiff™ Hematopoietic Kit. At the end of the differentiation period, cells were harvested from the supernatant and analyzed by flow cytometry for co-expression of CD34+ and CD45+ . (A) Representative density plots showing CD34+ and CD45+ expression, (B) percentage of cells co-expressing CD34+ and CD45+ , and (C) total number of viable cells harvested are shown. Data are expressed as the mean (± SEM); n=4.
Protocols and Documentation
Find supporting information and directions for use in the Product Information Sheet or explore additional protocols below.
This product is designed for use in the following research area(s) as part of the highlighted workflow stage(s). Explore these workflows to learn more about the other products we offer to support each research area.
Resources and Publications
Educational Materials (16)
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STEMdiff™ Hematopoietic Kit
PRODUCTS ARE FOR RESEARCH USE ONLY AND NOT INTENDED FOR HUMAN OR ANIMAL DIAGNOSTIC OR THERAPEUTIC USES UNLESS OTHERWISE STATED. FOR ADDITIONAL INFORMATION ON QUALITY AT STEMCELL, REFER TO WWW.STEMCELL.COM/COMPLIANCE.