ROIs extraction from breast tissues is key to differentiating malignant from benign lesions. ROIs are the areas where tumors that grow with angiogenesis support have higher metabolic rates and produce more deoxyhemoglobin than normal tissues. Since deoxyhemoglobin is sensitive to light at 640 nm, it generally appears as a dark blue light-absorbing region in the image [34 (link)]. Therefore, we consider dark blue areas in a series of sequential DOBI data as ROIs or SAs and design an algorithm to extract these ROIs. Detailed ROIs extraction processes are provided in the Supplement 1.