The syngeneic orthotopic murine bladder cancer model MB49 is hampered by unreliable tumor implantation. We optimized this model by a simple modification of the standard implantation technique in three groups of mice. Fifty thousand (group I), 20,000 (group II), or 10,000 (group III) tumor cells were implanted into cauterized bladders by transurethral instillation, and dwell time was prolonged to 3 h. Tumor take, survival, and bladder weights were determined as outcome variables. To verify whether this modification maintained its sensitivity to topical immunotherapy, an initial tumor load of 100,000 MB49 cells was given, and mice were treated intravesically with Bacillus Calmette-Guérin or phosphate-buffered saline. The prolonged dwell time of tumor cells resulted in take rates of 100% in all three groups. Survival and bladder weights were significantly correlated with the number of instilled cells. Even with the highest tumor load, Bacillus Calmette-Guérin therapy improved survival and reduced bladder weights significantly, as compared to PBS. Thus, the modified model is highly reliable and maintains its susceptibility to topical immunotherapy.