The bladder cancer cell line BK-10 was established from a grade III-IV transitional cell carcinoma (TCC). BK-10 is near-tetraploid (+/-4n) and consists of two subclones with 20-25 structural aberrations. Here we report the cytogenetic analysis of BK-10 by G-banding, spectral karyotyping (SKY), and FISH. SKY refers to the hybridization of 24 differentially labeled chromosome painting probes and the simultaneous visualization of all human chromosomes using spectral imaging. SKY enabled us to confirm 12 markers in BK-10 previously described by G-banding, redefine 11 aberrations, and detect 4 hidden chromosomal rearrangements, 2 of which had been identified as normal or deleted copies of chromosome 20 and 1 as a normal chromosome 3. Twenty out of 21 translocations identified were unbalanced. FISH analysis of BK-10 using chromosome arm-specific paints, centromere probes, and oncogene/tumor suppressor gene-specific probes revealed a deletion of CDKN2A (p16) in all copies of chromosome 9, a low-level amplification of MYC (five copies), and loss of one copy of TP53; detected the presence of the Y chromosome in a hidden translocation; and detected four copies of ERBB-2. A probe set for BCR and ABL verified breakpoints for all translocations involving chromosomes 9 and 22. A new karyotype presentation, "SKY-gram," is introduced by combining data from G-banding, SKY, and FISH analysis. This study demonstrates the approach of combining molecular cytogenetic techniques to characterize fully the multiple complex chromosomal rearrangements found in the bladder cancer cell line BK-10, and to refine the chromosomal breakpoints for all translocations.