murine models of human carcinogenesis are exceedingly valuable tools to understand genetic mechanisms of neoplastic growth the identification of recurrent chromosomal rearrangements by cytogenetic techniques serves as an initial screening test for tumour specific aberrations in murine models of human carcinogenesis however karyotype analysis is technically demanding because mouse chromosomes are acrocentric and of similar size fluorescence in situ hybridization fish with mouse chromosome specific painting probes can complement conventional banding analysis although sensitive and specific fish analyses are restricted to the visualization of only a few mouse chromosomes at a time here we apply a novel imaging technique that we developed recently for the visualization of human chromosomes to the simultaneous discernment of all mouse chromosomes the approach is based on spectral imaging to measure chromosome specific spectra after fish with differentially labelled mouse chromosome painting probes utilizing a combination of fourier spectroscopy ccd imaging and conventional optical microscopy spectral imaging allows simultaneous measurement of the fluorescence emission spectrum at all sample points a spectrum based classification algorithm has been adapted to karyotype mouse chromosomes we have applied spectral karyotyping sky to chemically induced plasmocytomas mammary gland tumours from transgenic mice overexpressing the c myc oncogene and thymomas from mice deficient for the ataxia telangiectasia atm gene results from these analyses demonstrate the potential of sky to identify complex chromosomal aberrations in mouse models of human carcinogenesis.