Cancer is currently a common problem facing human beings. Early diagnosis is a key factor in reducing cancer mortality. Due to its sensitivity to the microenvironment, fluorescence lifetime imaging microscopy (FLIM) can not only distinguish early cancer tissues from normal tissues, but also monitor the drug treatment of cancer. Therefore, FLIM has great application potential in cancer diagnosis. This article introduces the basic principles and detection methods of FLIM, summarizes the endogenous and exogenous fluorescence characteristics and their relationship with cancer. In addition, this article mainly reviews the recent applications of FLIM in the diagnosis of cancers in the nervous system, respiratory system, digestive system, reproductive system, urinary system, endocrine system and skin system, and discusses the application advantages, limitations and future development trends of FLIM in cancer diagnosis.