Fig. 2

Essential hPSC characteristics were maintained throughout culture on recombinant laminin-521 (LN-521) in Essential 8™ Flex Medium (E8), shown for hESC1. a Typical undifferentiated colony morphology the day after passaging (upper picture, scale bar = 50 μm) and a higher magnification of the cells at confluency prior to the next passaging (day 4, lower picture, scale bar = 20 μm). Cells at feeder-free passage level 4, passage 40 in total. b Expression of pluripotency markers NANOG, OCT-3/4, SSEA-3, SSEA-4, TRA-1-60, and TRA-1-81, and no expression of early differentiation marker SSEA-1, at passages 8–13. Nuclei counterstained with DAPI; scale bars = 200 μm. c Flow cytometry analysis of human embryonic stem cells (hESCs) cultured in feeder-free conditions compared to hESCs cultured on hFF feeder cells in standard hESC medium. Blue histogram for positive, black for negative (unstained) sample, and red for isotype control. d Expression of markers of the three embryonic germ layers after spontaneous differentiation. Nuclei counterstained with DAPI; scale bars = 200 μm. e Cells after 25 passages showing normal female karyotype in KaryoLite BoBs assay. The results are shown as signal relative to karyotypically normal female (/F, red) and male (/M, blue) genomic DNA used as a reference (equal to 1) for each of the 24 chromosomal probes (cover both p and q arms of all chromosomes). Software threshold for changes shown as a green line and deviating results in red