Figure 3 | Scientific Reports

Figure 3

From: Generation of 3D retinal tissue from human pluripotent stem cells using a directed small molecule-based serum-free microwell platform

Figure 3

Retinal differentiation in 3D AMM using animal-derived supplements (xeno). (A) Schematic of the 3D AMM xeno small molecule protocol (B) Bright-field microscopy of aggregates generated from MShef10 ESC line in AMM at day 0 (first image from left) and differentiated 3D optic vesicle-like structures at day 60 of differentiation (scale bars 500 µm) (C) Histological and immunohistological analysis of CRX and PAX6 in retinal organoids generated from MShef10 ESC line at day 42 of differentiation. (D) RT PCR analysis of relative expression of retinal-specific markers (mean ± SD; n = 3 independently cultured AMMs each containing over 50 OVs from N = 1 ESC line, MShef10. (E,F) Histological and immunohistological analysis of CRX, RCVRN, L/M OPSIN, RHO and PKCa at week 32 of differentiation in retinal organoids generated from MShef10 ESC line. 3D retinal organoids which were transferred from AMM into suspension culture from week 6 of differentiation grew larger and maintained laminated organisation and exhibited longer outer segment-like protrusion beyond outer limiting membrane (F) compared to organoids which were maintained in AMM throughout the study (E).

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