A method for nucleic acid hybridization to isolated chromosomes in suspension.
A procedure was developed to provide differential fluorescent staining of metaphase chromosomes in suspension following nucleic acid hybridization. For this purpose metaphase chromosomes were isolated from a Chinese hamster X human hybrid cell line. After hybridization with biotinylated human genomic DNA, the human chromosomes were visualized by indirect immunofluorescence using antibodies against biotin and fluoresceine-isothiocyanate-(FITC)-labeled second antibodies. This resulted in green fluorescent human chromosomes. In contrast, Chinese hamster chromosomes revealed red fluorescent staining only when counterstained with propidium iodide. Notably, interspecies chromosomal rearrangements could be easily detected. After hybridization and fluorescent staining, chromosomes still showed a well-preserved morphology under the light microscope. We suggest that this procedure may have a useful application in flow cytometry and sorting.
Dudin
G
G
Cremer
T
T
Schardin
M
M
Hausmann
M
M
Bier
F
F
Cremer
C
C
D000818:Animals; D002877:Chromosomes, Human; D006224:Cricetinae; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D006801:Humans; D006822:Hybrid Cells; D007621:Karyotyping / Q000379:methods*; D009693:Nucleic Acid Hybridization
D005456:Fluorescent Dyes