Two cancer cell lines, MCF7 and ISHIKAWA, were each obtained from two sources, which maintained cells in different culture media. MCF7 was derived from breast tumor: #1 was obtained from ATCC at passage 149 and passaged an additional 16 times; #2 was cultured in another laboratory for unknown passages and subcultured an additional 95 times. Cell line #2 had a baseline HPRT mutation frequency 10.fold higher than #1. ISHIKAWA was derived from an endometrial tumor: #3 was obtained from European Collection of Cell Cultures passaged more than 3 times, was ER positive, and subcultured 130 more times; #4 was subcultured unknown times in another laboratory, was ER negative, and subcultured an additional 132 times. Cell line #4 had a baseline HPRT mutation frequency 1400-fold higher than #3. MCF7 cultures had complex karyotypes, however, similarities included up to 83 chromosomes. additional chromosomes 2,3,4,6,6,7,8.9,10,12,13,14,15,17,19,20, structural abnormalities add(3)(p25),del(6) (q21),add(22)(q13), and at least 1 of up to 17 marker chromosomes in common. The ISHIKAWA cultures each had up to 60 chromosomes with 4-6 markers, but only a missing X in common. CGH studies were performed using different colored fluorochromes to label each of the two MCF7 or ISHIKAWA cell lines, which were then co-hybridized to normal mataphases. Differences seen between the pairs of MCF7 and ISHIKAWA cultures reflect karyotype differences. Some Initial DNA polymorphic data for the ISHIKAWA cell lines (Dr. R. Bamezai, New Delhi, India) suggest that they may not be derived from the same established cell line. Our studios demonstrate the utilization of CGH for comparing cell lines originating from the same specimen, but undergoing karyotypic and mutation rate changes due to different culture conditions and passage numbers.