[Uniparametric analysis of chromosomes from human fibroblasts using flow cytofluorometry].
Metaphase chromosomes from human diploid fibroblasts were isolated by the polyamine method and stained with different fluorochromes: propidium iodide, chromomycine A2, bisbensimide H33258, and DAPI. The fluorescently stained chromosomes were analyzed using a cell sorter ATC-3000 ("Bruker-Spectrospine"), and the flow histograms were investigated to identify the peaks of individual chromosomes and to choose the chromosomes which can be isolated to a high purity. High resolution was obtained with H33258 or DAPI. It was demonstrated that the peaks of chromosomes 13, 17, 20 are well distinguished from the adjacent peaks, so that these chromosomes can be sorted with a high purity. The peaks formed by chromosomes 5, 6, 8, 14, 15 are slightly overlapped by the adjacent peaks, and the sorting on the basis of monovariate analysis cannot provide sufficient purity of the sorted fractions (< 0.5). Chromosomes 1-2, 3-4, 7-X, 9-12, 16-18, 19, 21-22 cannot be discriminated for successful sorting of an individual chromosome type. The data obtained provided an experimental basis for choosing the chromosomes to be sorted with high purity for the subsequent cloning of their DNA.
Nasedkina
T V
TV
Poltorykhin
S A
SA
Slezinger
S I
SI
Poletaev
A I
AI
D002478:Cells, Cultured; D002877:Chromosomes, Human; D005347:Fibroblasts / Q000378:metabolism; D005434:Flow Cytometry; D005456:Fluorescent Dyes; D006801:Humans
D005456:Fluorescent Dyes
Odnoparamtericheskiĭ analiz khromosom iz fibroblastov cheloveka metodom tsitofluorimetrii v potoke.