Calcein am staining solution

The cells were assayed for growth activity in the hydrogel on days 1, 3, and 5. The complete medium was discarded to add PBS to wash the cells 2–3 times. In the dark, the prepared Calcein−AM staining solution (Beyotime, Shanghai, China) 100 µL was added to each well and incubated at 37 °C for 25 min. The pictures of stained cells were recorded using a fluorescence microscope.

8505C cells were seeded into a 35‐ mm glass bottom culture dish. The cells were resuspended in PBS and loaded with calcein AM staining solution (Beyotime Biotechnology, NO. C2009S) and 3 × CoCl₂ in the dark at 37°C for 30–45 min. At the end of incubation, culture medium was replaced with medium preheated to 37°C, and the dishes were incubated at 37°C for 30 min in order to ensure that calcein AM was hydrolyzed by lactamase to produce green fluorescent calcein. The cells were washed with PBS 2–3 times, and the fluorescence image of mitochondrial calcein were acquired using a Leica TCS SP8 MP laser scanning confocal microscope.