Generating high-purity cardiac and endothelial derivatives from patterned mesoderm using human pluripotent stem cells

DOI: 10.1038/nprot.2016.153

Authors: Nathan J Palpant; Lil Pabon; Clayton E Friedman; Meredith Roberts; Brandon Hadland; Rebecca J Zaunbrecher; Irwin Bernstein; Ying Zheng; Charles E Murry

Step 1: iPSC/ESC maintainance

day 0 ~ 1

culture “adherent” human PSC in “mTeSR” for “1” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Step 2: Primitive streak 1

day 1 ~ 2

culture “adherent” human PSC in “RPMI” for “1” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Step 3: Primitive streak 2

day 2 ~ 3

culture “adherent” human PSC in “RPMI” for “1” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “1” days.

Step 4: endothelial 1

day 3 ~ 6

culture “adherent” human PSC in “StemPro” for “3” days. For ECM, use “Matrigel” at “N/A” “N/A”. Medium should contain factors shown in the following table, and should be changed once in “3” days.

Ideally, “EC” should express “KDR”, “CD34”, “PECAM1”, “CDH5” at the end of culture.

Step 5: endothelial 2

day 6 ~ 11

culture “adherent” unknown species cell in “EGM” for “5” days. For ECM, use “Gelatin” at “0.1” “%”. Medium should contain factors shown in the following table, and should be changed once in “5” days.

Ideally, “EC” should express “PECAM1”, “CDH5” at the end of culture.