Health Information

Separation of large chromosomes of eukaryotes is not possible by conventional electrophoresis. Fluorescence-activated cell sorting, also known as flow cytometry or flow karyotyping, can separate the individual chromosomes of eukaryotes.

Fluorescence-activated cell sorting

To carry out FACS, the dividing cells are carefully broken open, and a mixture of intact chromosomes is prepared. These chromosomes are then stained with a fluorescent dye. The quantity of the dye that binds to a chromosome depends on its size. Thus, larger chromosomes bind more dye and fluoresce more brightly than the smaller ones.

The dye-mixed chromosomes are diluted and passed through a fine aperture that results in the formation of a stream of droplets. Each droplet contains a single chromosome. A laser detects the fluorescence of the chromosomes. When the fluorescence indicates that the chromosome illuminated by the laser is the one desired, that does not contain the desired chromosome pass through a waste collection vessel.

Collection of chromosomes with identical size

The direct application of FACS is not suitable for the separation of chromosomes with identical sizes. Chromosomes 21 and 22 in humans can be achieved by use of special dyes (Hoechst 33258 and chromomycin A3), which bind to AT-rich DNA, or GC-rich DNA.

It is convenient to separate two or more chromosomes with identical sizes by different dyes. This is possible since no two chromosomes are likely to contain identical GCC/AT contents.