Figure 1.

Morphological evaluation and biochemical confirmation of human colon organoids from cryo-preserved frozen samples. (A) Recovered human colon APC^−/−;KRAS^G12D organoids from frozen stocks. Frozen vials of organoids were thawed at 37°C, re-suspended in BME gel, and plated into a 24-well plate as a 50-µl droplet/well. The medium was refreshed every 3‒4 days. The growth of organoids was monitored with the ImageXpress^micro automated-imaging system (10× objective). (B) Robust growth of human colon APC^−/−;KRAS^G12D organoids (4× objective). Representative images from two different wells of a 24-well plate are shown. All the images shown are merged images of Z-stack using ImageXpress software. (C) The KRAS^G12D protein expression of human colon APC^−/−;KRAS^G12D organoids was confirmed in comparison to that of APC^−/−;KRAS^WT organoids. Organoids were collected for western blotting analysis with anti-KRAS^G12D-specific antibody.