1 FISH Techniques, FISH Probes and Their Applications in Medicine and Biology - An Overview
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probes in a single-color approach. Figure 4A shows an example of a normal analysis using the centromere-specific probe for chromosome X, and
in Fig. 4B the result of a whole chromosome painting using a probe for
chromosome 8 is shown.
When complex chromosomal rearrangements are identified by conventional banding techniques, it is possible to get a clue as to which chromosomes are involved and by using a multi-color ratio-labeling approach
it later became possible to characterize the chromosomal rearrangement
in greater detail (Fig. 4C).
The so-called microdeletion syndromes are an increasingly important
group of specific genetic disorders that are characterized by discrete but
consistent chromosomal deletions initially identified by high-resolution
banding methods (prophase techniques). These methods have an estimated resolution of 2-5 megabase pairs which is barely sufficient to demonstrate such microdeletions, and at the same time these methods are
labor-intensive as well as technically difficult. Therefore, even with high
quality banding, this type of syndrome can only be identified in approximately 50% of the cases (Dobyns et al. 1991). FISH analyses with suitable
cosmid or YAC clones, on the other hand, are ideal for the demonstration
of these microdeletion syndromes. The first microdeletion syndrome to be
diagnosed with FISH was Miller-Dieker syndrome (Kuwano et al. 1991),
which is a rare malformation syndrome, manifested by lissencephaly
..
Fig. 4. Different examples of FISH techniques used for identification of specific chromosomal aberrations. A Detection of highly repeated sequences. Staining of alpha-satellite
DNA of chromosome X using cloned alpha-satellite probe labeled indirectly with FITC.
Counterstained with propidium iodine. B CISS hybridization with a chromosome 8 specific
probe indirectly labeled with FITC, propidium iodine used as counterstain. Image captured
using a confocal microscope. C Three-color ratio labeling hybridization with libraries for
chromosomes 2 (red), 8 (green) and 15 (yellow - 50% red and 50% green) identifying a
complex translocation that cannot be completely diagnosed by high resolution banding
(upper row). D Locus-specific hybridization at chromosome region 22ql1.2 (red) and a control signal (green) for identification of the microdeletion syndrome DiGeorge. E Dual color
arm-specific hybridization with Xp (red) and Xq (green) showing that Q-banding only identifies a structural abnormality in Xp of unknown origin, but FISH identifies a partial trisomy
for Xq and a partial monosomy for Xp. F Interphase FISH hybridization identifying an extra
chromosome 18 centromere (green) in the nuclei (blue). G Telomeric probe hybridization
for chromosome 3 (p-arm green and q-arm red) showing two normal chromosome 3,
whereas the small marker chromosome (from Fig. SF) does not contain chromosome 3derived telomeric DNA. H In situ hybridization to permeabilized cell nuclei identifying
cytoplasmic mRNA coding for medium chain acyl coenzyme A dehydrogenase (MCAD)
(green). Details on experimental conditions can be obtained by contacting the authors.
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probes in a single-color approach. Figure 4A shows an example of a normal analysis using the centromere-specific probe for chromosome X, and
in Fig. 4B the result of a whole chromosome painting using a probe for
chromosome 8 is shown.
When complex chromosomal rearrangements are identified by conventional banding techniques, it is possible to get a clue as to which chromosomes are involved and by using a multi-color ratio-labeling approach
it later became possible to characterize the chromosomal rearrangement
in greater detail (Fig. 4C).
The so-called microdeletion syndromes are an increasingly important
group of specific genetic disorders that are characterized by discrete but
consistent chromosomal deletions initially identified by high-resolution
banding methods (prophase techniques). These methods have an estimated resolution of 2-5 megabase pairs which is barely sufficient to demonstrate such microdeletions, and at the same time these methods are
labor-intensive as well as technically difficult. Therefore, even with high
quality banding, this type of syndrome can only be identified in approximately 50% of the cases (Dobyns et al. 1991). FISH analyses with suitable
cosmid or YAC clones, on the other hand, are ideal for the demonstration
of these microdeletion syndromes. The first microdeletion syndrome to be
diagnosed with FISH was Miller-Dieker syndrome (Kuwano et al. 1991),
which is a rare malformation syndrome, manifested by lissencephaly
..
Fig. 4. Different examples of FISH techniques used for identification of specific chromosomal aberrations. A Detection of highly repeated sequences. Staining of alpha-satellite
DNA of chromosome X using cloned alpha-satellite probe labeled indirectly with FITC.
Counterstained with propidium iodine. B CISS hybridization with a chromosome 8 specific
probe indirectly labeled with FITC, propidium iodine used as counterstain. Image captured
using a confocal microscope. C Three-color ratio labeling hybridization with libraries for
chromosomes 2 (red), 8 (green) and 15 (yellow - 50% red and 50% green) identifying a
complex translocation that cannot be completely diagnosed by high resolution banding
(upper row). D Locus-specific hybridization at chromosome region 22ql1.2 (red) and a control signal (green) for identification of the microdeletion syndrome DiGeorge. E Dual color
arm-specific hybridization with Xp (red) and Xq (green) showing that Q-banding only identifies a structural abnormality in Xp of unknown origin, but FISH identifies a partial trisomy
for Xq and a partial monosomy for Xp. F Interphase FISH hybridization identifying an extra
chromosome 18 centromere (green) in the nuclei (blue). G Telomeric probe hybridization
for chromosome 3 (p-arm green and q-arm red) showing two normal chromosome 3,
whereas the small marker chromosome (from Fig. SF) does not contain chromosome 3derived telomeric DNA. H In situ hybridization to permeabilized cell nuclei identifying
cytoplasmic mRNA coding for medium chain acyl coenzyme A dehydrogenase (MCAD)
(green). Details on experimental conditions can be obtained by contacting the authors.