Figure 2.
Establishment of Stable Transgenic ArcLight-hiPSCs
(A) Expression of ArcLight (green fluorescence) by undifferentiated ArcLight-hiPSC colonies (top), but not by the parental hiPSC colonies (bottom). The merged light (DIC) and fluorescent images (right) are also shown. Scale bars, 50 μm.
(B) Immunostaining of ArcLight-hiPSC colonies for OCT4, NANOG, SSEA4, and TRA-1-60.
(C) Immunostaining of ArcLight-hiPSC-CMs for sarcomeric α-actinin and cTnI.
(D) qPCR analysis of ArcLight-hiPSC-CMs showing expression of cardiac-specific genes (NKX2.5, MLC-2V, MYH-6, and MYH-7) and downregulation of pluripotent genes (OCT4 and NANOG). Experiments included three biological replicates measured as two technical replicates.
(E and F) ArcLight expression by hiPSC-CMs monolayers (E) and single-dispersed cells (F) and the derived optical AP. Note the increase in fluorescence during repolarization (resting state) and the decreased intensity during depolarization (AP development). Scale bar, 20 μm.
(G) Optical tracings analysis used to measure APD90. This process can be scaled up to analyze several cells (bottom).
Error bars represent SEM.
See also Figure S2.