I2959, widely known as Irgacure 2959, is a photoinitiator extensively used for ultraviolet light-induced photocrosslinking of hydrogels in biomedical applications and is particularly utilized in combination with carboxymethylcellulose (CMA) hydrogels as a bioink for bio 3D printing applications; furthermore, I2959 is recognized for its high radical generation efficiency under UV exposure but also exhibits concentration-dependent cytotoxicity to rapidly dividing cell lines.

Encapsulation of Saos-2 cells in photochemically cross-linked CMA hydrogels:
1. Proliferate the human osteosarcoma cell line Saos-2 in RPMI medium (containing 15% FBS, 1% L-glutamine, and 1% penicillin/streptomycin).
2. Resuspend the cells in a neutralized CMA solution supplemented with 0.02% or 0.1% photoinitiator I2959.
3. Add 45 µL of the cell suspension (approximately 5,000 cells) to each well of a 96-well plate and incubate at 37°C for 30 minutes to allow for initial gelation.
4. Irradiate the hydrogels with 365 nm UV light for either 1 minute or 10 minutes to complete photocrosslinking; a control hydrogel without UV exposure was also prepared.
5. Culture the encapsulated cell hydrogels in α-MEM medium (supplemented with 50 µg/mL ascorbic acid, 10 mM β-glycerophosphate, 10% FBS, and 1% penicillin/streptomycin) for up to 14 days.