Purpose A new way in breast cancer detection, Dynamic Optical Breast Imaging (DOBI) was studied. Materials and Methods 52 patients receiving breast biopsy and DOBI were enrolled in this study. 19 were proved of breast cancer, 33 were benign. Results 94.8% of blue area in non-breast cancer lesions was found as wandering or diffusive pattern, while 68.42% breast cancer showed focal pattern. 86.46% of the curve signature of blue area in non-breast cancer lesions was wavy or flat, while 57.37% of breast cancer showed a steep decline. In 64.58% non-breast cancer, the curve of blue area was similar to that of non-blue area. 78.95% breast cancer had their curve different from that of non-blue area. The absolute value of amplitude (-5.77±2.13) of blue area in cancer was higher than that in non-cancer ( -3.34±0.87). The differences were all statistically significant(P<0.05). Conclusion The spatial and temporal characteristics of DOBI were of diagnostic and differential value for breast cancer. The absolute value of amplitude, over|-5|, also helped the diagnosis of breast cancer.