Chapter 6
In Situ Hybridization for DNA:
Fluorescent Probe
MORIMASA MATSUTA AND MAYUMI MATSUTA
Introduction
Fluorescence in situ hybridization (FISH) with chromosomespecific probes is one of the non-isotopic in situ hybridization
methods that opens up several new areas of cytogenetic investigation by allowing visual determination of the presence and
normality of specific genetic sequences in single metaphase or
interphase cells (Pinkel 1986a, 1986b). With this approach,
the genetic loci to be analyzed are made microscopically visible
in single cells using in situ hybridization with nucleic acid probes
specific to these loci. The DNA in target cells is made as a single
strand and incubated with single-stranded fluorochromedirectly-labeled or chemically modified DNA probe under conditions where the probe will anneal only with DNA sequences to
which it has high DNA sequence homology. The bound chemically modified probe is made visible by immunofluorescent detection methods. This technique is often called standard FISH.
Multi-target FISH (M-FISH) provides the means of detecting
multiple genetic targets simultaneously within individual nuclei
or metaphase spreads. By combining probes for a number of targets into a single hybridization, precious specimens are conserved, assay time and labor are reduced, and the clonal or mosaic nature of a specimen can be accurately assayed (VYSIONS
1997).
A recently described M -FISH technology, 24-color FISH using
spectral karyotyping (SKY), permits the simultaneous visualizaCorrespondence to Morimasa Matsuta, Iwate Medical University, Department of Obstetrics and Gynecology, 19-1 Uchimaru, Morioka, 020-8505,
Japan (phone +81-19-651-5111 extension 2344; fax +81-19-622-1900;
e-mail matsutao@iwate-med.ac.jp)
In Situ Hybridization for DNA:
Fluorescent Probe
MORIMASA MATSUTA AND MAYUMI MATSUTA
Introduction
Fluorescence in situ hybridization (FISH) with chromosomespecific probes is one of the non-isotopic in situ hybridization
methods that opens up several new areas of cytogenetic investigation by allowing visual determination of the presence and
normality of specific genetic sequences in single metaphase or
interphase cells (Pinkel 1986a, 1986b). With this approach,
the genetic loci to be analyzed are made microscopically visible
in single cells using in situ hybridization with nucleic acid probes
specific to these loci. The DNA in target cells is made as a single
strand and incubated with single-stranded fluorochromedirectly-labeled or chemically modified DNA probe under conditions where the probe will anneal only with DNA sequences to
which it has high DNA sequence homology. The bound chemically modified probe is made visible by immunofluorescent detection methods. This technique is often called standard FISH.
Multi-target FISH (M-FISH) provides the means of detecting
multiple genetic targets simultaneously within individual nuclei
or metaphase spreads. By combining probes for a number of targets into a single hybridization, precious specimens are conserved, assay time and labor are reduced, and the clonal or mosaic nature of a specimen can be accurately assayed (VYSIONS
1997).
A recently described M -FISH technology, 24-color FISH using
spectral karyotyping (SKY), permits the simultaneous visualizaCorrespondence to Morimasa Matsuta, Iwate Medical University, Department of Obstetrics and Gynecology, 19-1 Uchimaru, Morioka, 020-8505,
Japan (phone +81-19-651-5111 extension 2344; fax +81-19-622-1900;
e-mail matsutao@iwate-med.ac.jp)