This paper describes a fluorescence in situ hybridization (FISH) protocol to rapidly determine homozygosity of transgenic mice. Fibroblast nuclei from 4- to 6-week-old congenic progeny were hybridized with a human APP bacterial artificial chromosome (BAC) probe. Scoring of greater than 50 interphase nuclei demonstrated the presence of either one or two hybridization signals in more than 95% of the interphase nuclei from hemizygous and homozygous animals, respectively. Fibroblast nuclei from 4- to 6-week-old hybrid progeny were hybridized with a mouse BAC probe from the distal portion of mouse chromosome 16. More than 50 interphase nuclei were scored for hybridization signals. There were either 2 or 3 hybridization signals in more than 80% of the nuclei from diploid and Ts65Dn animals, respectively. The advantages of this FISH method are the relative ease of culturing fibroblasts, the presence of both interphase and metaphase nuclei and the qualitative nature of the fluorescent signal that eliminates the necessity of progeny testing.