Chapter 12
Chromosome Analysis by Fluorescence in situ
Hybrid ization
MARTIN BENTZ, STEFAN JOOS AND PETER LICHTER
lntroduction
In 1969 visualization of nucleic acid sequences in situ was first described
using isotopically labeled rRNA (Gall and Pardue 1969). Since then, in situ
hybridization employing radioactively labeled probes has been widely used.
With the advent of nonisotopic procedures for labeHing nucleic acids in the
early 1980s, fluorescence in situ hybridization (FISH) was developed and
has become increasingly popular. Compared with radioactive in situ hybridization, FISH has a number of specific advantages including increases in
speed, sensitivity and resolution (for review see Raap et al. 1990; Lichterand
Ward 1990; Trask 1991).
In situ hybridization permits the detection of single-copy nucleic acid
sequences in biological and clinical specimens. Using a suitable probe, entire chromosomes or chromosomal subregions may be visualized permitting detection of alterations in copy number or structure (translocations,
etc.). This can be achieved using metaphase chromosome spreads or interphase nuclei ("interphase cytogenetics"), examples for which are shown in
Figure 1 and 2 (Joseph et al. 1984; Cremer et al. 1986). FISHis based on the
specific base pairing of sequences of the DNA probe and complementary
sequences in cells of the specimen. The DNA probes are chemically modified by the introduction of reporter molecules (e.g. biotin or digoxigenin)
and are detected using fluorochromes conjugated to specific reporter-binding molecules ( as avidin or antiborlies directed against the reporter molecules). Alternatively, the probes can be labeled directlywith fluorochromes.
Martin Bentz, Univ. Heidelberg, Medizinische Klinik und Poliklinik V, Hospitalstr. 3,
Heidelberg, 69115, Germany, Stefan Joos, Deutsches Krebsforschungszentrum, Abt. Organisation komplexer Genome, Im Neuenheimer Feld 280, Heidelberg, 69120, Germany,
Correspondence to Peter Lichter, Deutsches Krebsforschungszentrum, Abt. Organisation komplexer Genome, Im N euenheimer Feld 280, Heidelberg, 69120, Germany (phone
+49-6221-424609; fax +49-6221-424639; e-mail p.lichter@dkfz-heidelberg.de)
Chromosome Analysis by Fluorescence in situ
Hybrid ization
MARTIN BENTZ, STEFAN JOOS AND PETER LICHTER
lntroduction
In 1969 visualization of nucleic acid sequences in situ was first described
using isotopically labeled rRNA (Gall and Pardue 1969). Since then, in situ
hybridization employing radioactively labeled probes has been widely used.
With the advent of nonisotopic procedures for labeHing nucleic acids in the
early 1980s, fluorescence in situ hybridization (FISH) was developed and
has become increasingly popular. Compared with radioactive in situ hybridization, FISH has a number of specific advantages including increases in
speed, sensitivity and resolution (for review see Raap et al. 1990; Lichterand
Ward 1990; Trask 1991).
In situ hybridization permits the detection of single-copy nucleic acid
sequences in biological and clinical specimens. Using a suitable probe, entire chromosomes or chromosomal subregions may be visualized permitting detection of alterations in copy number or structure (translocations,
etc.). This can be achieved using metaphase chromosome spreads or interphase nuclei ("interphase cytogenetics"), examples for which are shown in
Figure 1 and 2 (Joseph et al. 1984; Cremer et al. 1986). FISHis based on the
specific base pairing of sequences of the DNA probe and complementary
sequences in cells of the specimen. The DNA probes are chemically modified by the introduction of reporter molecules (e.g. biotin or digoxigenin)
and are detected using fluorochromes conjugated to specific reporter-binding molecules ( as avidin or antiborlies directed against the reporter molecules). Alternatively, the probes can be labeled directlywith fluorochromes.
Martin Bentz, Univ. Heidelberg, Medizinische Klinik und Poliklinik V, Hospitalstr. 3,
Heidelberg, 69115, Germany, Stefan Joos, Deutsches Krebsforschungszentrum, Abt. Organisation komplexer Genome, Im Neuenheimer Feld 280, Heidelberg, 69120, Germany,
Correspondence to Peter Lichter, Deutsches Krebsforschungszentrum, Abt. Organisation komplexer Genome, Im N euenheimer Feld 280, Heidelberg, 69120, Germany (phone
+49-6221-424609; fax +49-6221-424639; e-mail p.lichter@dkfz-heidelberg.de)