Cancer is a serious threat to human health, and there is an urgent need to develop new treatment methods to overcome it. Organelle targeting therapy, as a highly effective and less toxic side effect treatment strategy, has great research significance and development prospects. Being an essential organelle, the Golgi apparatus plays a particularly major role in the growth of cancer cells. Acting as an indispensable and highly expressed antioxidant in cancer cells, glutathione (GSH) also contributes greatly during the Golgi oxidative stress. Therefore, it counts for much to track the changes of GSH concentration in Golgi for monitoring the occurrence and development of tumor cells, and exploring Golgi-targeted therapy is also extremely important for effective treatment of cancer. In this work, we designed and synthesized a simple Golgi-targeting fluorescent probe GT-GSH for accurately detecting GSH. The probe GT-GSH reacting with GSH decomposes toxic substances to Golgi, thereby killing cancer cells. At the same time, the ratiometric fluorescent probe can detect the concentration changes of GSH in Golgi stress with high sensitivity and selectivity in living cells. Therefore, such a GSH-responsive fluorescent probe with a Golgi-targeted therapy effect gives a new method for accurate treatment of cancer.