To generate defined, serum-free EHM, purified cardiomyocytes were mixed with HFFs (American Type Culture Collection) at a 70%:30% ratio. The cell mixture was reconstituted in a mixture of pH-neutralized medical-grade bovine collagen (0.4 mg per EHM; LLC Collagen Solutions) and concentrated serum-free medium [2× RPMI, 8% B27 without insulin, penicillin (200 U/ml), and streptomycin (200 μg/ml)] and cultured for 3 days in Iscove medium with 4% B27 without insulin, 1% nonessential amino acids, 2 mM glutamine, 300 μM ascorbic acid, IGF1 (100 ng/ml; AF-100-11), FGF-2 (10 ng/ml; AF-100-18B), VEGF165 (5 ng/ml; AF-100-20), TGF-b1 (5 ng/ml; AF-100-21C; all growth factors are from PeproTech), penicillin (100 U/ml), and streptomycin (100 μg/ml) (32 (link)). After a 3-day condensation period, EHM were transferred to flexible holders to support auxotonic contractions. Analysis was carried out after a total EHM culture period of 4 weeks.
Protocol detail
Defined, Serum-Free Engineered Heart Muscle
Partial Protocol Preview
This section provides a glimpse into the protocol.
The remaining content is hidden due to licensing restrictions, but the full text is available at the following link:
Access Free Full Text.
Amino acids
Ascorbic acid
Bovine
Cardiomyocytes
Cell
Collagen
Fgf 2
Glutamine
Growth factors
Igf1
Insulin
Penicillin
Serum
Streptomycin
Tgf b1
Vegf165
Top 5 similar protocols
Variable analysis
independent variables
- Ratio of purified cardiomyocytes to HFFs (70%:30%)
- Analysis of engineered heart muscle (EHM) after a total culture period of 4 weeks
- PH-neutralized medical-grade bovine collagen (0.4 mg per EHM)
- Concentrated serum-free medium [2× RPMI, 8% B27 without insulin, penicillin (200 U/ml), and streptomycin (200 μg/ml)]
- Iscove medium with 4% B27 without insulin, 1% nonessential amino acids, 2 mM glutamine, 300 μM ascorbic acid, IGF1 (100 ng/ml), FGF-2 (10 ng/ml), VEGF165 (5 ng/ml), TGF-b1 (5 ng/ml), penicillin (100 U/ml), and streptomycin (100 μg/ml)
- 3-day condensation period before transferring EHM to flexible holders
- Positive control: Not explicitly mentioned
- Negative control: Not explicitly mentioned
Annotations
Based on most similar protocols
Etiam vel ipsum. Morbi facilisis vestibulum nisl. Praesent cursus laoreet felis. Integer adipiscing pretium orci. Nulla facilisi. Quisque posuere bibendum purus. Nulla quam mauris, cursus eget, convallis ac, molestie non, enim. Aliquam congue. Quisque sagittis nonummy sapien. Proin molestie sem vitae urna. Maecenas lorem.
Sign up for free or login to display all annotations
As authors may omit details in methods from publication, our AI will look for missing critical information across the 5 most similar protocols.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!