Fluorescence in Situ Hybridization – FISH
Fluorescence in situ hybridization (FISH) is the molecular cytogenetic technique
that allows cytogeneticists to analyze chromosome resolution at the DNA or gene
level. FISH can be performed on dividing (metaphase) and non-dividing
(interphase) cells to identify numerical and structural abnormalities resulting from
genetic disorders.
In FISH, cytogeneticists utilize one or more FISH probes that typically fall into one
of the following three categories:
1. Repetitive sequences, including alpha satellite DNA, that bind to the
centromere of a chromosome;
2. DNA segments, representative of the entire chromosome, that will bind to
and cover the entire length of a particular chromosome; and
3. DNA segments from specific genes or regions on a chromosome that have
been previously mapped or identified.
A probe is "tagged" directly, by incorporating fluorescent nucleotides, or
indirectly, by incorporating nucleotides with attached small molecules, such as
biotin, digoxygenin, or dinitrophenyl, to which fluorescent antibodies can later be
bound. The probe and the chromosomes that are being analyzed are denatured
and allowed to bind or hybridize to one another. If necessary, antibodies with a
fluorescent tag are applied to the cells. The cells are then viewed with a
fluorescence microscope. The fluorescent signals represent the probe that is
bound to the chromosomes.