Fig. 3

ON2 showed equal or superior efficiency compared to common xeno-free culture media. a 253G1 hiPSC clone morphology on iMatrix-511 with E8, AK02N, or ON2, respectively, over 20 passages. Scale bar, 200 µm. b Doubling time comparison of 253G1 hiPSCs maintained in different xeno-free culture media: E8, AK02N, or ON2. (Passage number = 20, **p < 0.01, n.s., no significant difference; data are mean ± SD). c Flow cytometric analysis of SSEA4, TRA-1-60, and SSEA1. Gray solid histograms show the isotype control populations, and color hollow histograms show the stained populations. The percentages of marker-positive cells are presented in each graph. Cell events were normalized to the mode. d Immunofluorescence staining of pluripotency markers in hiPSCs: Oct3/4 (green); Nanog (red). Nuclei were stained with DAPI dye (blue). Scale bar, 100 µm. e qRT-PCR analysis of pluripotent gene expression levels of OCT4, NANOG, SOX2, and KLF4 in 253G1 hiPSCs in three different xeno-free culture media. (n = 6 independent samples, and data are mean ± SD, *p < 0.05, ** or ^##p < 0.01, n.s. no significant difference)