Urovysiont fluorescence in situ hybridization (FISH) is a sensitive and specific test used to diagnose urothelial carcinoma in urine. It detects aneuploidy of chromosomes 3, 7 and 17, and loss of both 9p21 loci in malignant urothelial cells. We evaluated Urovysiont FISH in non-urothelial carcinoma involving bladder to determine its possible application to their diagnosis and surveillance. Paraffin blocks from 31 non-urothelial bladder carcinomas, 12 pure urothelial carcinomas and 2 urothelial carcinomas with squamous differentiation were tested according to Vysis-Abbot Laboratories' recommended standards. Cases included 15 primary squamous carcinoma, 2 urothelial carcinoma with squamous differentiation, 4 primary adenocarcinoma, 5 colonic, 4 prostatic and 1 cervical adenocarcinoma. Total 60% of squamous, 83% of pure urothelial, 100% of urothelial carcinoma with squamous differentiation and 100% of primary and secondary adenocarcinomas hybridized successfully; 2/10 (11%) squamous carcinomas and 11/14 (79%) primary and secondary adenocarcinomas were Urovysiont FISH-positive with primary adenocarcinomas accounting for 75% (3/4), colonic, 80% (4/5), prostatic, 75% (3/4) and cervical, 100% (1/1) positivity. Total 70% (7/10) of pure urothelial carcinomas and 100% (2/2) of urothelial carcinomas with squamous differentiation were Urovysiont FISH-positive. In conclusion, we found that chromosomal abnormalities tested for by Urovysiont FISH may be seen in non-urothelial carcinomas of bladder. These false-positive results were frequent in primary and secondary adenocarcinoma and rare in squamous carcinoma. This has significant implications for the accurate diagnosis and management of patients with urinary tract cancer. Urovysiont FISH cannot be used to definitively diagnose squamous carcinoma or adenocarcinoma nor can it be used to differentiate the two from urothelial carcinoma. However, it may be useful as a surveillance tool in established primary and secondary bladder adenocarcinoma. Cytopathologists and urologists should correlate Urovysiont FISH results with cytomorphology and clinical information.