Hydrogel have been a great material for biomedical applications. In this study, fluorescent hydrogels formulated from carrageenan biopolymer and firefly luciferin, exhibited high fluorescence quantum yield and stability, thermo-reversibility, and high transparency. Importantly, the hydrogels are pH-responsive allowing them to be used as sensing materials for a range of pH from 3 to 6.5. The hydrogels may find potential applications as cheap and tailored materials in important fields such as pH sensor for wound healing or bioluminescence hydrogels for cell study. Graphical abstract: [Figure not available: see fulltext.].