Journal: Developmental Cell

Article Title: Inducible Stem-Cell-Derived Embryos Capture Mouse Morphogenetic Events In Vitro

doi: 10.1016/j.devcel.2020.12.004

Figure Lengend Snippet: iETX Embryos Express Canonical Post-implantation Embryo Markers (A) Top: Time course and schematic of iETX embryo generation from cell seeding on day 0 to day 5. Bottom: Images of a representative iETX embryos at specific time points. Scale bar, 100 μm. (B) The contents of a single AggreWell were collected at day 4 to quantify formation efficiency of iETX embryos. ES compartment and VE-like layer express CAG-GFP; the TS compartment is unlabeled. Scale bar, 200 μm. Panels (right) highlight representative structures such as well-formed iETX embryos (1 and 2); an inflated iETX embryo (3); a well-formed iETX embryo in the process of inflating (4). Scale bar, 150 μm. (C) Quantification of iETX embryo formation at either 3 or 4 days. Day 3 = 1,542/4,187 structures (n = 4); day 4 = 960/4,410 structures (n = 4). Error bar: SD. (D) Time-lapse still images of iETX embryo formation over the course of 67 h. Top: The ES compartment and VE-like layer express CAG-GFP; TS compartment is unlabeled. Bottom, CAG-GFP ESCs alone (gray). White arrowheads highlight TS cells, yellow arrow and asterisk the forming ES lumen. Six examples. Scale bar, 30 μm. (E) iETX embryos at 1 day of development. ES and TS compartment are enclosed with a dashed line. Dox-treated CAG-tetOG4 ESCs transiently express mCherry and downregulate CAG-GFP (arrows). 91/109 structures from 2 independent experiments. Scale bar, 30 μm. (F–I) iETX embryo at day 4 stained for (F) Gata6 (green), Oct4 (gray), and Eomes (red) (41/42 structures). (G) Gata4 (green), Oct4 (gray), and Ap2γ (red) (18/19 structures). (H) iETX embryo at day 4 stained for Sox17 (green), Oct4 (gray), and Cdx2 (red) (21/21). (I) iETX embryo at day 3 stained for Otx2 (purple), Eomes (green), and DAPI (blue) (27/31). (F–H) n = 3 each; scale, 30 μm. (I) n = 3; scale bar, 20 μm. (J) Lineage quantification in iETX embryos in comparison with E5.5 and ETX embryos. 15 iETX, n = 3; 10 ETX and 10 E5.5 embryos, n = 3: from ( Sozen et al., 2018 ). Error bars: SD. ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗∗ p < 0.0001, ns, nonsignificant. See also Figure S2 .

Article Snippet: To prepare the AggreWell plate (STEMCELL Technologies 34415), 500μl of anti-adherence rinsing solution (STEMCELL Technologies 07010) was added to each well.

Techniques: Staining, Comparison