The Petri Dish-N2B27 Culture Condition Maintains RPE Phenotype by Inhibiting Cell Proliferation and mTOR Activation

Figure 1

Sequential dissection of DA rat retina to obtain primary RPE cells using dispase. (a) Eyes were dissected out from DA rats, and extraocular muscle attachments (black arrow) were removed from the eye under the dissecting microscope. (b) After incubation in the 2% dispase solution, eyes were washed in PBS. Then the retina/RPE complex (blue arrow) was isolated by removing the choroid (green arrow), the anterior cornea (red arrow), and lens (black arrow). The RPE monolayer was still attached to the retina. (c, d) The resulting retina/RPE complex was cut to small pieces and was further incubated in 10% FBS containing medium for 20 min at 37°C to allow the RPE (blue arrow) to detach from the neural retina (yellow arrow). (e) The detached RPE sheets were collected and digested in 0.1% trypsin solution and then gently pipetted up and down several times to achieve smaller patches of RPE cells. (f) Primary RPE cells of DA rats after 24 h on cell culture dish show high confluence and pigmentation. Scale bar: 100 μm.

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