STEMdiff™ Ventricular Cardiomyocyte Differentiation Kit
Serum-free media for differentiation of human PSCs to ventricular cardiomyocytes and long-term maintenance of human PSC-derived cardiomyocytes
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- Gentle Cell Dissociation Reagent
cGMP, enzyme-free cell dissociation reagent
- Y-27632 (Dihydrochloride)
RHO/ROCK pathway inhibitor; Inhibits ROCK1 and ROCK2
- D-PBS (Without Ca++ and Mg++)
Dulbecco’s phosphate-buffered saline without calcium and magnesium
STEMdiff™ Cardiomyocyte Maintenance Kit (Catalog #05020) comprises the maintenance basal medium and supplement; it can be used for long-term maintenance of hPSC-derived cardiomyocytes for one month or longer. These cardiomyocytes can be used in various downstream applications and analyses.
NOTE: This product was formerly named ‘STEMdiff™ Cardiomyocyte Differentiation Kit’; the product itself and manufacturing procedures have not changed, but the name has been updated to more accurately reflect the cell type generated.
Figure 1. Cardiomyocyte Differentiation Protocol
Two days before the differentiation protocol, hPSC colonies are harvested and seeded as single cells at 350,000 cells/well in a 12-well format in TeSR™ medium. After one day (Day -1), the medium is replaced with fresh TeSR™ medium. The following day (Day 0), the TeSR™ medium is replaced with Medium A (STEMdiff™ Cardiomyocyte Differentiation Basal Medium containing Supplement A) to begin inducing the cells toward a cardiomyocyte fate. On day 2, a full medium change is performed with fresh Medium B (STEMdiff™ Cardiomyocyte Differentiation Basal Medium containing Supplement B). On days 4 and 6, full medium changes are performed with fresh Medium C (STEMdiff™ Cardiomyocyte Differentiation Basal Medium containing Supplement C). On day 8, medium is switched to STEMdiff™ Cardiomyocyte Maintenance Medium with full medium changes on days 10, 12 and 14, to promote further differentiation into cardiomyocyte cells. Small beating areas of cardiomyocytes can be seen as early as day 8, progressing to a full lawn of beating cardiomyocytes that can be harvested as early as day 15.
Figure 2. Morphology of hPSC-Derived Cardiomyocytes
Representative images of (A) hES (H9) cells and (B) hiPS (WLS-1C) cells on day 15 of differentiation to cardiomyocytes using the STEMdiff™ Ventricular Cardiomyocyte Differentiation Kit. Differentiated cells exhibit typical cardiomyocyte morphology as an adherent, tightly packed web-like monolayer of beating cells. (C) Representative confocal microscopy image of a single hPSC-derived cardiomyocyte generated with the STEMdiff™ Ventricular Cardiomyocyte Differentiation Kit and stained with cTnT (green) and DAPI (blue).
Figure 3. Efficient and Robust Generation of cTnT-Positive Cardiomyocytes
hES and hiPS cells were cultured for 15 days in single wells of 12-well plates using the STEMdiff™ Ventricular Cardiomyocyte Differentiation Kit. At the end of the culture period, cells were harvested and analyzed by flow cytometry for expression of cardiac troponin T (cTnT). (A) Histogram analysis for cardiomyocyte cell marker cTnT for cultures of hES (H9) and hiPS (WLS-1C and STiPS-M001) cells. (Filled = sample; blank = secondary antibody only control) (B,C) Percentages and total numbers of cells expressing cTnT in cultures of hES or hiPS cells are shown. Data shown as mean ± SEM; n=3.
Figure 4. hPSC-Derived Cardiomyocytes Exhibit a Robust and Stable Excitability Profile
Microelectrode array (MEA) voltage recordings of cardiomyocytes (day 27) derived from human pluripotent stem cells generated and maintained with the STEMdiff™ Cardiomyocyte Differentiation and Maintenance Kits. The hPSC-derived cardiomyocytes have a characteristic electrical profile and stable beat rate. A large depolarization spike followed by a smaller repolarization deflection is observed.
Figure 5. Generation of Cardiomyocytes from hPSCs Maintained in mTeSR™ Plus
Human ES (H9) and iPS (WLS-1C) cells were maintained in mTeSR™1 (daily feeds) or mTeSR™ Plus (restricted feeds) and differentiated to cardiomyocytes using the STEMdiff™ Ventricular Cardiomyocyte Differentiation Kit. At the end of the differentiation period, cells were harvested and analyzed by microelectrode array (MEA) and flow cytometry. (A) Representative MEA voltage recordings of cardiomyocytes (day 20) demonstrate a characteristic electrical profile and stable beat rate. (B) Percentages of cells expressing cTNT and (C) total number of viable cells harvested are shown. Data are expressed as the mean (± SEM); n=2.
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 (11)
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STEMdiff™ Ventricular Cardiomyocyte Differentiation 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.