Objective To study the role of fibulin-5 in urothelial carcinoma of bladder. Methods Fibulin-5 expression was detected in bladder cancer tissues (13 cases of G_1 and G_2, 7 cases of G_3) and normal bladder mucosa samples by Western blotting assay. Fibulin-5 cDNA was amplified by PCR and cloned into pMD-19T simple vector. The pMD-19T-Fibulin-5 vector was digested by restriction endonucleases XhoI and EcoRI to generate a XhoI-Fibulin5-EcoRI fragment that was then ligated into the identical sites in p-EGFP-Nl plasmid to synthesize p-EGFP-Fibulin-5 plasmid. The p-EGFP-Fibulin-5 plasmid was finally transfected into bladder cancer cell line 5637. The migration and invasion of untransfected, vector-transfected and fibulin-5-transfected bladder cancer cells were measured by Boyden chamber assay. Results Compared to 1. 16 ±0. 28 in the normal control, the expression of fibulin-5 protein in low grade and high grade tumors were 0. 57±0. 32 and 0. 44±0. 42(P<0. 01, respectively). However, the difference between low grade and high grade tumors was not statistically significant (P>0. 05). The successfully transfected bladder cancer cells demonstrated green fluorescent light. The migrated cell number of fibulin-5-transfected cells was 127. 6 ± 3. 1 compared with 139. 3±7. 7 for vector-transfected cells and 136. 9±5. 7 for untransfected cells (P>0. 05, respectively). In contrast, the invaded cell number of fibulin-5-transfected cells was 8. 0±3. 1 compared with 31. 5±4. 8 for vector-transfected cells and 31. 7±4. 7 for untransfected cells (P<0. 01, respectively). Conclusion Fibulin-5 is down-regulated in urothelial carcinoma of bladder and acts as a tumor suppressor gene by inhibiting the invasion of bladder cancer cells.