Cardiac Myocyte Differentiation Protocol

Direct CM differentiation was carried as previously described.^{5 (link),29 (link),30 (link)} Briefly, after cell expansion on Plastic MC, cells/MC aggregates were re-plated on LN521-coated plates. The cells were treated with 12 μM Gsk3 inhibitor CHIR99021 (Selleck) in RPMI/B27 without insulin (Life Technologies) for 24 h, followed by a treatment with 5 μM inhibitor of Wnt production-2 (IWP2; Stemgent) at day 3. The medium was replaced on day 5 by fresh RPMI/B27 without insulin and cells were then maintained in this medium until day 10, when they were maintained in RPMI/B27 with insulin (B27^® Supplement; Life Technologies) until day 15. The differentiated cells were then trypsinized into a single-cell suspension and fixed with 4% paraformaldehyde, followed by staining with 1:40 diluted anti-Cardiac myosin heavy chain (MF20; Developmental Studies Hybridoma Bank) and 1:200 anti-troponin 1 cardiac (cTnT; Millipore) before FACS analysis, as described earlier.

Free full text: Click here

Lam A.T., Li J., Chen A.K., Birch W.R., Reuveny S, & Oh S.K. (2015). Improved Human Pluripotent Stem Cell Attachment and Spreading on Xeno-Free Laminin-521-Coated Microcarriers Results in Efficient Growth in Agitated Cultures. BioResearch Open Access, 4(1), 242-257.

Publication 2015

Gsk3 inhibitor CHIR99021 RPMI/B27 without insulin RPMI/B27 B27® Supplement

Cardiac Cardiac myosin Cell Chir99021 Gsk3 Hybridoma Insulin Paraformaldehyde Supplement Troponin 1

Corresponding Organization :

Other organizations : Agency for Science, Technology and Research

Top 5 similar protocols

Variable analysis

independent variables

Treatment with 12 μM Gsk3 inhibitor CHIR99021 for 24 h
Treatment with 5 μM inhibitor of Wnt production-2 (IWP2) at day 3

dependent variables

Percentage of cells positive for cardiac myosin heavy chain (MF20) and troponin 1 cardiac (cTnT) as measured by FACS analysis

control variables

Cell expansion on Plastic MC
Re-plating cells/MC aggregates on LN521-coated plates
Maintenance in RPMI/B27 without insulin medium until day 10, then in RPMI/B27 with insulin medium until day 15

controls

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.

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?

Revolutionizing how scientists
search and build protocols!