The mechanism and principles of autofluorescence imaging based on autofluorescence technique are reported. The threshold value of fluorescence spectrum ratio applied can be quantitative and objective and the reliable measurement method that may provide intuitive method of autofluorescence imaging in the gut mucosa. The suspected lesion may be found rapidly according to the imaging color difference, therefore the results of clinical study of the digestive tract cancer diagnosis indicated that the sensitivity, specificity, and diagnostic accuracy were 94%, 95.5% and 94.8% respectively, and it has very high value in clinical application.