Optical imaging has been widely used in basic research and clinical follow-up as an emerging in vivo imaging technology. It can provide real-time physiological and pathological information that cannot be obtained from in vitro testing. When compared to traditional imaging strategies, NIR optical imaging offers advantages such as spatiotemporal resolution and a relatively high signal-to-noise ratio. This review is primarily concerned with the structure design of molecular NIR fluorescent probes and their bioimaging and diagnosis applications in typical nephron-urological diseases, such as chronic kidney disease (CKD), acute kidney injury (AKI), iatrogenic ureteral injury (IUI), kidney neoplasms, and bladder cancer (BC). Additionally, the potential development and current difficulties in this field are also presented in this review.