Background: Fibroblast growth factor 21 (FGF21), a secreted protein, plays a crucial role in regulating metabolism and energy homeostasis. Nevertheless, the expression pattern of Fgf21 across diverse tissues and its responsiveness to various dietary regimens remain incompletely understood. Methods: In this study, we developed a Fgf21-enhanced green fluorescent protein (EGFP) reporter mouse model to explore the expression of endogenous Fgf21 in different tissues under four dietary conditions: normal chow, low-protein diet, fasting, and fasting-refeeding. Results: A low-protein diet was found to induce Fgf21 expression in both the liver and skeletal muscle. Notably, Fgf21 was predominantly expressed in the periportal region of the liver. In the pancreas, Fgf21 exhibited a patchy expression pattern in the exocrine portion, but was absent in the endocrine part, regardless of the dietary regimens. Regarding the spleen, fasting triggered the expression of Fgf21, which was mainly localized in the red pulp area. Moreover, under fasting conditions, Fgf21 showed a scattered expression pattern in the small intestine. Conclusions: The Fgf21-EGFP reporter mouse model serves as a valuable tool for dissecting the expression of endogenous Fgf21 in different tissues under various dietary and stress conditions. Further investigations using this model may contribute to uncovering the hitherto unrecognized functions of locally produced FGF21.