Two-photon excitation fluorescence lifetime imaging is an ingenious technique which can provide both morphological and biochemical information of tissues in three-dimensional space with subcellular resolution. The structural and functional properties of tissue can be quantitatively characterized in detail. Therefore, this technique provides a powerful tool for the noninvasive, label-free, intravital imaging of biological tissues, and which is promising to be applied in clinical diagnosis of tumors. In recent years, this technology has been used to detect tumors, and which has already become one of the research foci in the field of biomedicine. In this review paper, firstly, the concept of the two-photon excitation fluorescence lifetime and common fluorescence lifetime detection methods are introduced simply; secondly, we summarize the recent progresses of using two-photon excitation fluorescence lifetime imaging to detect various tumors, including digestive tract tumors, brain tumors, skin tumors, and so on, and among them, our results on the detection of gastric cancers and glioma are elaborately depicted; Finally, the potential advantages and possible challenges of this technology in future clinical application are provided and discussed.