Murine embryonic stem (ES) cells were cultured on a material containing immobilized leukemia inhibitory factor (LIF). To immobilize LIF, we synthesized photoreactive gelatin mixed with LIF and cast the mixture on a polystyrene plate, which was then dried. LIF was immobilized by photoirradiation in the presence or absence of a photo mask. The plate was washed until LIF was no longer released. Murine ES cells were cultured on the immobilized LIF. Activation of STAT3 was maintained on the immobilized LIF for 6 d even after removing soluble LIF. Oct-3/4 was also expressed in the cells cultured on the immobilized LIF. As a result, the mouse ES cells were cultured without differentiating on the immobilized LIF for 6 d. It was possible to culture murine ES cells without adding soluble LIF at each medium change. We conclude that our material containing immobilized LIF might be useful in the culture of murine ES cells.