Kit Summary
Reagents for the directed differentiation of human pluripotent stem cells into the cardiomyocyte lineage.
Key Benefits
- Optimized reagents generate more robust and uniform beating of differentiated cardiomyocytes.
- Provides consistent differentiation across pluripotent stem cell lines
- Reproducible differentiation protocols translate into cost and time savings
- Maximizes workflow efficiency by standardizing cardiomyocyte differentiation
- Applicable for drug toxicity and small molecule screening
Why Use Pre-mixed Differentiation Cocktails when Differentiating Human Pluripotent Stem Cells into Cardiomyocytes?
The StemXVivo® Cardiomyocyte Differentiation Kit uses high-quality specialized media and pre-mixed differentiation cocktails to maximize differentiation efficiency and ensure the consistent and reliable generation of scalable amounts of cardiomyocytes. Using optimized reagents and a straightforward protocol, this kit provides a reproducible method for obtaining high-yields of healthy cardiomyocytes while minimizing the cost and time involved in the differentiation process. This kit has been tested on iPSC cell lines derived from both blood and fibroblasts.
Cardiomyocyte differentiation in vitro:
- Uses premixed differentiation cocktails to optimally drive reproducible differentiation of pluripotent stem cells into cardiomyocytes.
- Yields a highly enriched and healthy cardiomyocyte population.
- Produces cardiomyocytes that express Cardiac Troponin T and contract synchronously.
- Can be part of small molecule and drug toxicity screening workflows.
Precautions
The acute and chronic effects of overexposure to reagents of this kit are unknown. Safe laboratory procedures should be followed and protective clothing should be worn when handling kit reagents.
Kit Contents
This kit contains the following reagents to drive pluripotent stem cell differentiation into cardiomyocytes and an antibody to verify differentiation status.
- Stem Cell Qualified RGF BME, Pathclear®
- Cardiomyocyte Differentiation Base Media Supplement I
- Cardiomyocyte Differentiation Base Media Supplement II
- Cardiomyocyte Differentiation Cocktail I
- Cardiomyocyte Differentiation Cocktail IIA
- Cardiomyocyte Differentiation Cocktail IIB
- Cardiomyocyte Differentiation Cocktail III
- Anti-Human Cardiac Troponin T Antibody
The quantity of each component in this kit is sufficient to differentiate two 24-well plates, or an equivalent surface area, of pluripotent stem cells into cardiomyocytes.
Stability and Storage
Store unopened kit at < -70 °C in a manual defrost freezer. Do not use past kit expiration date.
Limitations
- FOR LABORATORY RESEARCH USE ONLY. NOT FOR DIAGNOSTIC USE.
- Do not mix or substitute reagents with those from other lots or sources.
- The safety and efficacy of this product in diagnostic or other clinical uses has not been established.
- The quality and differentiation potential of human pluripotent stem cells at the onset of the differentiation protocol are of paramount importance to the efficiency of differentiation.
Data Examples
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Synchronized Contractile Beating of iPSC-Derived Cardiomyocytes.
Cardiomyocytes were differentiated from JOY1 human induced pluripotent stem cells using the StemXVivo® Cardiomyocyte Differentiation Kit. Waves of beating cardiomyocytes were visualized using brightfield microscopy.
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iPSC-Derived Cardiomyocyte Contractions Visualized Using the Calcium Indicator, Fluo-4.
Cardiomyocytes were differentiated from JOY1 human induced pluripotent stem cells using the reagents and protocol provided in this kit. They were assessed for their ability to contract using the Fluo-4 calcium binding assay.
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Differentiation of Pluripotent Stem Cells into Cardiomyocytes.
JOY6 (left panel) and iBJ6 (right panel) human induced pluripotent stem cells were differentiated into cardiomyocytes using the media supplements included in this kit. Aside from visually observing contracting cells, commitment to the cardiomyocyte cell fate was evaluated by labeling with the Anti-Human Cardiac Troponin T antibody included in this kit. For visualization, the cells were stained using NorthernLights™ 557-conjugated Donkey anti-Mouse Secondary Antibody (R&D Systems, Catalog # NL007; red), and the nuclei were counterstained with DAPI (blue).
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Improved Efficiency of iPSC Differentiation Across Pluripotent Stem Cell Lines.
Multiple established human iPS cell lines and one embryonic stem cell (ESC) line were differentiated into beating cardiomyocytes using either the new and enhanced version of the StemXVivo® Cardiomyocyte Differentiation Kit (New Kit) or the previous version of the kit (SC032). After 21 days of differentiation beating quality was scored based on percentage of cells beating and beating intensity. Differentiation with the new kit results in more robust and uniform differentiation based on beat quality. The new kit also increases the likelihood of differentiation success across ES and iPS cell lines.
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Endothelin-1 Induces Expression of Atrial Natriuretic Peptide in Cardiomyoctyes Derived from iPSCs Using the New Kit.
JOY6 human iPSCs were differentiated into beating cardiomyocytes using the new and enhanced version of the StemXVivo® Cardiomyocyte Differentiation Kit. After 21 days of differentiation cardiomyocytes were treated with 5 nM of Endothelin-1 (Catalog # 1160) for 25.5 hours to induce cardiac hypertrophy. The level of secreted Pro-Atrial Natriuretic peptide (ANP), which is known to be elevated during cardiac hypertrophy, was assessed using (A) Western Blot and (B) ELISA (Human NTProANP Quantikine® ELISA; Catalog # DANP00). As expected, detection of Pro-ANP was increased in Endothelin-1 treated cells.
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Kit-Derived Cardiomyocytes Express Atrial and Ventricular Markers.
JOY6 human iPSCs were differentiated into cardiomyocytes using the new and enhanced version of the StemXVivo® Cardiomyocyte Differentiation Kit. The iPSC-derived cardiomyocytes were fixed and processed for immunocytochemical analysis of atrialspecific (MLC2a) and ventricle-specific (MYH7; Rabbit Anti-Human MYH7 Monoclonal Antibody; Catalog # MAB90961) marker expression.
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Kit-Derived Cardiomyocytes Express Stage-specific Markers During Differentiation.
JOY6 human iPSCs were differentiated with the new StemXVivo® Cardiomyocyte Differentiation Kit and assessed at select time points for stage-specific marker expression. The pluripotency marker Oct-4A (Mouse Anti-Human Oct-4A Monoclonal Antibody; Catalog # MAB17591) is highly expressed during early differentiation (Day 0) and is subsequently downregulated. Expression of the mesoderm marker, Snail (Goat-Anti-Human Snail Polyclonal Antibody; Catalog # AF3639), is expressed intermediately during differentiation (Day 1). The cardiomyocyte markers NKX2.5 (Goat Anti-Human NKX2.5 Polyclonal Antibody; Catalog # AF2444) and Troponin T (Mouse Anti-Human Cardiac Troponin T Monocolonal Antibody; Catalog # MAB1874) are not present in cells during early (Day 0) and intermediate (Day1) differentiation and become more highly expressed during the later stages of differentiation (Day 7, Day 30). Snail and NKX2.5 primary antibodies were visualized with the NorthernLights™ (NL)557-conjugated Donkey Anti-Goat IgG Secondary Antibody (Catalog # NL001). Oct-4A and Troponin T were visualized with the NL557-conjugated Donkey Anti-Mouse IgG Secondary Antibody (Catalog # NL007).
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