1022-7954/02/3802- $27.00 © 2002
Russian Journal of Genetics, Vol. 38, No. 2, 2002, pp. 95–104. Translated from Genetika, Vol. 38, No. 2, 2002, pp. 149–160.
Original Russian Text Copyright © 2002 by Ostroverkhova, Nazarenko, Cheremnykh.
Genomic imbalance in cells of different organisms
is usually detected by classical cytogenetic and molec-
ular genetic analyses. Comparative genomic hybridiza-
tion (CGH) is a relatively new technique, which has
assumed a great importance in the past several years.
By using the CGH assay, genomic changes can be
revealed by comparison the DNA copy number in cells
of test and control tissues in a single experiment [1–5].
The CGH assay was ﬁrst described by Kallioniemi
in 1992 . The principle and basic steps of the CGH
analysis are shown in the ﬁgure. To label the test and
reference DNA, the two different reporter molecules
are used, for example, biotin and digoxigenin. After the
incorporation of the reporter molecules, a mixture of
test and reference DNAs (1 : 1) is hybridized to normal
metaphase chromosomes of a healthy individual in the
presence of an excess of unlabeled repetitive Cot1 DNA
added to suppress the repeated sequences. Upon two-
color ﬂuorescent microscopy, the reporter molecules
are identiﬁed by using some ﬂuorochrome-labeled
agents exhibiting high afﬁnity to the former. For exam-
ple, biotin-labeled test DNA can be detected by using
ﬂuorescein isothiocyanate-labeled avidin exhibiting
high afﬁnity to biotin, whereas digoxigenin-labeled ref-
erence DNA can be revealed by using antidigoxigenin
antibodies conjugated with rhodamin. The two reagents
display different ﬂuorescence, green and red, respec-
tively. In the DNA sample examined, chromosome
imbalance is detected from difference in intensity of
green and red ﬂuorescence, i.e., from ﬂuorescence ratio
(FR) of two ﬂourochromes along each chromosome.
FR values correspond to the relative content of the
detected DNA sequences as compared to the control.
The correspondent FR proﬁles are obtained by quanti-
tative digital image analysis of green and red ﬂuores-
cence intensity along each chromosome.
In the absence of chromosome imbalance between
tested and reference DNAs, green to red ﬂuorescence
ratio, FR, is equal to 1.0. A duplication of a chromo-
some region or trisomy for some chromosome leads to
higher FR value of the tested tissue, whereas chromo-
some deletions or monosomy results in lower FR on the
corresponding proﬁles. For example, at complete or
partial trisomy of a deﬁnite chromosome, the average
green-to-red FR is about 1.5, whereas at monosomy it
is equal to 0.5. In the case of 50% mosaic karyotype,
the expected FR is 1.25 for the trisomic clone and 0.75
for the monosomic one.
Note that only relative
changes in the DNA copy number as compared to the
control can be revealed by the CGH analysis [1–3].
The CGH analysis proved to be very helpful in
examining solid tumor cell karyotype, because in this
case, the adequate chromosome preparations are difﬁ-
cult to obtain by conventional cytogenetic methods [4–
12]. In tumor cells, the high level of DNA ampliﬁcation
is often observed, which can be visualized by ﬂuores-
cent microscopy as an intense signal from the corre-
spondent chromosome regions. However, the quantita-
tive hybridization analysis is of great importance when
the low-copy-number gains and losses of genetic mate-
rial are present in the test tissue or when chromosomal
aberration occur in a few cells. In these cases, an
approach was used when determining the FR proﬁle
along each chromosome was followed by averaging of
individual proﬁle from several metaphase spreads to
study the deviations of speciﬁc chromosome segments
from the normal values . In addition to identiﬁca-
tion of speciﬁc genomic gains and losses in tumor cells,
the CGH analysis proved to be helpful in studying the
Comparative Genomic Hybridization As a New Method
for Detection of Genomic Imbalance
N. B. Ostroverkhova, S. A. Nazarenko, and A. D. Cheremnykh
Research Institute of Medical Genetics, Tomsk Research Center, Russian Academy of Medical Sciences, Tomsk, 634050 Russia;
fax: (3822) 51-37-44; e-mail: firstname.lastname@example.org
Received May 5, 2001; in ﬁnal form, July 30, 2001
—Comparative Genomic Hybridization (CGH) is a molecular cytogenetic method for detecting chro-
mosomal imbalances by comparing the copy number of DNA sequences in cells of tested tissue and the refer-
ence specimen. CGH is based on two-color ﬂuorescence suppressive in situ hybridization of genomic test and
reference DNAs, each labeled with a different ﬂuorochrome, to metaphase chromosomes of a healthy individual.
First described by Kallioniemi
in 1992, the CGH assay has been widely used for identiﬁcation and char-
acterization of both numerical and unbalanced structural chromosome abnormalities in cells of different tissues
at various pathological conditions in humans, especially in tumor diseases. We discuss the speciﬁc features and
quality control of comparative genomic hybridization, its advantages and limitations in detection of genomic
imbalance and the prospects for development of this technology.