Bladder cancer (BC) is a common human malignancy. Conventional ultrasound and white light cystoscopy are often used for BC diagnosis and resection, but insufficient specificity results in a high BC recurrence rate. New strategy for the diagnosis and resection of BC are needed. In this study, we developed a highly specific peptide-based probe for BC photoacoustic imaging (PAI) diagnosis and near-infrared (NIR)-imaging-guided resection post-instillation. A BC-specific peptide (PLSWT7) was selected by in vivo phage display technology and labeled with IRDye800CW to synthesize a BC-specific dual-modality imaging (DMI) probe (PLSWT7-DMI). The feasibility of PLSWT7-DMI-based dual-modality PAI-NIR imaging was assessed in vitro, in mouse models, and ex vivo human bladders. An air-pouch BC (APBC) model suitable for probe instillation was established to evaluate the probe-based BC PAI diagnosis and NIR-imaging-guided resection. Human bladders were used to assess whether PLSWT7-DMI-based DMI strategy is a translatable approach for BC detection and resection. The probe exhibited excellent selectivity and specificity both in vitro and in vivo. Post-instillation of the probe, tumors <3 mm were detectable by PAI, and NIR-imaging-guided tumor resection decreased the BC recurrence rate by 90% and increased the survival in the mouse model. Additionally, ex vivo NIR imaging of human bladders indicated that PLSWT7-DMI-based imaging would potentially allowed precise resection of BC in clinical settings. This PLSWT7-DMI-based DMI strategy was a translatable approach for BC diagnosis and resection and could potentially lower the BC recurrence rate.