Bioluminescence imaging (BLI) has been applied in gene therapy and research to screen for transgene expression, progression of infection, tumor growth and metastasis, and transplantation. It enables real-time and relatively noninvasive localization and serial quantification of biological processes in experimental animals. In diabetes research, BLI has been employed for the quantification of β-cell mass, monitoring of islet graft survival after transplantation, and detection of reporter gene expression. Here, we explore the use of BLI in a transgenic mouse expressing luciferase under the control of the mouse insulin 1 promoter (MIP-Luc-VU). A previous report on MIP-Luc-VU mice showed luminescence intensities emitted from the islets correlated well with the number of islets in vitro and in vivo. In this study, we showed MIP-Luc-VU mice fed a high fat diet for 8 weeks gave rise to a greater bioluminescent signal than mice fed a regular diet for the same period of time. Conversely, there was a strong reduction in the signal observed in diabetic Mafa-deficient/Mafk-transgenic mutant mice and streptozotocin-treated mice, reflecting the loss of β-cells. Furthermore, we were able to monitor fetal β-cell genesis in MIP-Luc-VU mice during the late gestational stage in a noninvasive and repetitive manner. In summary, we show that bioluminescence imaging of mice expressing a β-cell specific reporter allows detection of changes in β-cell mass and visualization of fetal β-cell neogenesis in uteri.