Counterstaining human chromosomes for flow karyology

Abstract
Isolated human metaphase chromosomes stained with the fluorochromes 4′-6-diamidino-2-phenylindole (DAPI) and chromomycin A3 (CA3), and counterstained with nonfluorescent netropsin (NTR), have been analyzed by dual-laser flow cytometry. Counterstaining with NTR reduces DAPI fluroscence except at regions on chromosomes 1,9,15,26, and Y, corresponding to C-band heterochromatin. Bivariate flow karyology of human chromosomes treated with this triple-stain combination resolves chromosomes 1,9, and Y distinctly from the remaining chromosomes and resolves variations between chrosome homologues not detected by staining with propidium iodide (PI) or with the double stain combination Hoechst 33258 (HO) and CA3.