Background: N-myc downstream regulated gene 1 (NDRG1) was involved in cell differentiation and was recently reported to exert various effects in tumorigenesis. The aim of this study was to assess its diagnostic value in urine as a useful marker for bladder cancer (BC). Methods: In this study, we recruited 119 BC patients, 65 patients with non-cancerous bladder diseases, and 60 healthy volunteers as control. Their urine concentrations of NDRG1, nuclear matrix protein 22 (NMP22), and creatinine (Cr) were measured and relevant clinical information was retrieved from their medical history records. Results: The expression of NDRG1/Cr and NMP22/Cr in urine were significantly higher in BC patients than those in non-cancerous bladder diseases (p = 0.009 and p = 0.023) and healthy controls (p = 0.005 and p = 0.002). The level of NDRG1/Cr was significantly associated with pathologic T stage (p < 0.001) and pathological grade (p < 0.001). The ROC of NDRG1/Cr to diagnose BC was 0.713 (95% CI, 0.630 - 0.797), with a sensitivity of 63.8% and a specificity of 73.4% at a cutoff of 76.3 ng/mg. NMP22/Cr was 0.705 (95% CI, 0.626 - 0.784), with a sensitivity of 64.2% and a specificity of 66.2% at a cutoff of 12.1 ng/mg. NDRG1/Cr in combination with NMP22/Cr shows a ROC of 0.719 (95% CI, 0.632 0.806) with a sensitivity of 64.9% and specificity of 75.9% Conclusions: Urine NDRG1 may be useful in a minimally invasive modality for determining bladder cancer. Predictive value of the two biomarkers was slightly higher than that of routine NMP22 parameter alone.