Improving the early detection rate and surveillance of bladder cancer remains a great challenge in medicine. Here, we identified sixteen proteins including Gc-globulin (GC) in urine from bladder cancer patients and normal controls by two-dimensional fluorescent differential gel electrophoresis (2D-DIGE) and matrix-assisted laser desorption time-of-flight mass spectrometry (MALDI-TOF/TOF MS). Bioinformatics analyses indicated GC played important roles in the regulation of growth, apoptosis, death and epidermal growth factor receptor activity. The GC expression patterns in urine or tissue from cases and controls were further quantified by western blotting, immunohistochemical staining and enzyme-linked immunosorbent assay (ELISA). ELISA quantification by correcting for creatinine expression showed GC-Cr was significantly increased in bladder cancer patients than in benign bladder damages cases and normal controls (1013.70+/-851.25 versus 99.34+/-55.87, 105.32+/-47.81ng/mg, respectively). Receiver operating characteristic (ROC) analysis suggested that at 161.086ng/mg urinary GC, bladder cancer could be detected with 92.31% sensitivity and 83.02% specificity, and 1407.481ng/mg with 82.61% sensitivity and 88.24% specificity could be used for the detection of infiltrating urothelial carcinoma of bladder cancer. Taken together, we identified GC as a potential novel urinary biomarker for the early detection and surveillance of bladder cancer.