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. Author manuscript; available in PMC: 2021 Apr 20.
Published in final edited form as: Methods Cell Biol. 2020 May 13;159:1–22. doi: 10.1016/bs.mcb.2020.04.009

FIG. 2.

FIG. 2

Protocol for the generation of human esophageal organoids. (A) hPSC are directly differentiated into definitive endoderm (day 3) and then into anterior foregut spheroids (day 6) which are then embedded in 3D matrix where differentiation and maturation of the spheroids continues for the generation of human esophageal organoids (HEOs). HEOs can survive up to 2 months in vitro, from day of embedding, by reducing organoids density 21 days after embedding (Day 27). Media and supplements are listed above each stage. For complete medium composition and supplements concentrations turn to Section 5.2 in the text. (B) Bright field images representing different stages of differentiation protocol. B-6 is zoom-in on dashed rectangle in B-5. Dashed circle in B-7 marks 3D matrix bubble margin. B-1, 5, 6, 7 and 8 were taken in up-right stereomicroscope (Leica, cat. # S8APO) scale bar: 500 μm. B-2, 3 and 4 were taken in inverted microscope (Nikon, TMS model), 10× magnification, scale bar—100 μm.