Stabilization of chromosomes by DNA intercalators for flow karyotyping and identification by banding of isolated chromosomes.
A number of structurally unrelated DNA intercalators have been studied as stabilizers of mitotic chromosomes during isolation from rodent and human metaphase cells. Seven out of the nine intercalators tested were found to be useful as chromosome stabilizing agents. Chromosome suspensions prepared in this way could be preserved for long periods of time. After isolation the chromosomal DNA was longer than 150 kb. With intercalated chromosomes high resolution flow karyotypes could be obtained as illustrated for the non-fluorescent intercalators 9-methylene-(1,3-dimethyl-2,4-dionepyrimidine-5-yl)-phenanthrid in iumchloride and 4'-aminomethyl-4,5', 8-trimethylpsoralen combined with DAPI and 33258 Hoeschst for fluorescent staining and for the fluorescent intercalator propidium iodide used as a stabilizer and as a fluorochrome. Passage of the intercalated chromosomes through the laser beam had no measurable effect on the length of the chromosomal DNA subsequently isolated. After flow analysis and collection on slides human chromosomes could easily be banded by Giemsa staining methods with the same resolution as obtained in conventional metaphase spreads. This allowed a ready identification of about 80 percent of all chromosomes in the unfractionated suspension collected after passage through the laser beam.
Aten
J A
JA
Laboratory for Radiobiology, University of Amsterdam, The Netherlands.
Buys
C H
CH
van der Veen
A Y
AY
Mesa
J R
JR
Yu
L C
LC
Gray
J W
JW
Osinga
J
J
Stap
J
J
D000818:Animals; D002460:Cell Line; D002871:Chromosome Banding; D002875:Chromosomes / Q000187:drug effects / Q000648:ultrastructure*; D002877:Chromosomes, Human / Q000187:drug effects / Q000648:ultrastructure*; D004247:DNA / Q000187:drug effects*; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007364:Intercalating Agents / Q000494:pharmacology*; D007621:Karyotyping
D007364:Intercalating Agents; D004247:DNA