Purpose SDF-1 is a ligand of the chemokine receptors CXCR4 and 7. The 6 known SDF-1 isoforms are generated by alternative mRNA splicing. While SDF-1 expression has been detected in various malignancies, only few groups have reported differential expression of SDF-1 isoforms and its clinical significance. We evaluated the expression of 3 SDF-1 isoforms (α, β and γ) in bladder cancer. Materials and Methods Using quantitative polymerase chain reaction we measured SDF-1α, β and γ mRNA levels in 25 normal and 44 bladder cancer tissues, and in 210 urine specimens (28 normal, 74 benign, 57 bladder cancer, 35 bladder cancer history, 8 other cancer history and 8 other cancer) from consecutive patients. Levels were correlated with clinical outcome. Results Of the SDF-1 isoforms only SDF-1β mRNA was significantly over expressed 2.5-fold to sixfold in bladder cancer compared to normal bladder tissues. SDF-1α was expressed in bladder tissues but SDF-1γ was undetectable. On multivariate analysis SDF-1β was an independent predictor of metastasis and disease specific mortality (p = 0.017 and 0.043, respectively). In exfoliated urothelial cells only SDF-1β mRNA levels were differentially expressed with 91.2% sensitivity and 73.8% specificity for detecting bladder cancer. In patients with a bladder cancer history increased SDF-1β levels indicated a 4.3-fold increased risk of recurrence within 6 months (p = 0.0001). Conclusions SDF-1 isoforms are differentially expressed in bladder tissues and exfoliated urothelial cells. SDF-1β mRNA levels in bladder cancer tissues predict a poor prognosis. Furthermore, SDF-1β mRNA levels in exfoliated cells detect bladder cancer with high sensitivity and they are a potential predictor of future recurrence.