Centromeric index versus DNA content flow karyotypes of human chromosomes measured by means of slit‐scan flow cytometry

We have applied slit‐scan flow cytometry (SSFCM) to classify human chromosomes according to their centromeric index (CI) and relative DNA content. The resulting bivariate—CI vs. DNA content—distributions shows 14 peaks for normal human chromosomes. Distinct peaks are produced by chromosomes 1, 2, 3, 4+5, 6+7+X, 8, 13+14+15, 16, 17+18, 19+20, and 21+22+Y. In addition, chromosomes 9 through 12 are resolved into three peaks. The identity of the chromosomes comprising each peak was determined by comparing CI vs. DNA content distributions measured for normal human chromosomes by means of SSFCM with CI and DNA content values measured for human chromosomes with image analysis. The accuracy of CI measurement by SSFCM was verified by measuring CIs for human chromosomes isolated from human/rodent hybrid cell lines containing only a few known human chromosomes. These studies showed CIs measured for human chromosomes 1–19 and 21 to be inclose agreement with the CIs calculated by means of image analysis. We further confirmed the chromosome assignments for each peak by showing that the relative volumes of the peaks in the CI vs. DNA content distributions for chromosomes from normal cells are similar to the relative frequencies of chromosomes expected for these peaks based on the peak assignments.