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Array-CGH, Karyotype Analysis, and FISH


The array-CGH test, which is already being used postnatally, will give obstetricians, geneticists, and their patients the opportunity in the prenatal setting to detect significantly more and smaller changes in the amount of chromosomal material present in individuals—and in significantly less time than a standard chromosome karyotype would take.

It may someday take the place of our standard techniques for cytogenetic analysis, but for now, it is a valuable addition to the available diagnostic tests.

Advances Over FISH

The technology, which has also been called chromosomal microarray, was first used to analyze gains and losses in chromosomal material in tumors and tumor cell lines. It is now a valuable tool in the postnatal testing of individuals with birth defects.

Between one-half and two-thirds of children with serious developmental abnormalities go undiagnosed and have a normal karyotype, so from a postnatal perspective, this new test has been welcomed at Johns Hopkins University and the Kennedy Krieger Institute, both in Baltimore, as well as at other institutions. Having a diagnosis facilitates the most appropriate therapy and allows parents to plan for future pregnancies and possible prenatal testing.

Yet it is the prenatal period for which array-CGH may have an even greater impact. Phenotypic features are not as apparent in the womb as at birth, making it more difficult to target testing with technology like rapid fluorescent in situ hybridization (FISH).

Along with standard karyotype analysis, the FISH technique has been the mainstay of cytogenetic analysis. It provides a targeted look at areas of the karyotype that are known to be associated with disease as a result of either the duplication or deletion of genetic material. In other words, it detects gains and losses in chromosomal material for just one or a few chromosome regions at a time.

Performing array-CGH is like doing FISH hundreds of times at once. Array-CGH testing may target the same chromosomal regions (and thus similar disorders) as a series of FISH tests, but array-CGH will target these regions at a much higher resolution, enabling the detection of much smaller deletions and duplications; it can also assess many regions associated with genetic disorders in a single test.

If we see on a prenatal ultrasound that a fetus has cardiac problems, for example, we might suspect the DiGeorge syndrome. The obstetrician today would probably perform an amniocentesis and order both a karyotype and FISH with a specific probe for the DiGeorge syndrome, which we know is caused by a deletion on chromosome 22, just as he or she would do in the postnatal period for a child with the syndrome's more obvious phenotypic features.

In the near future, the obstetrician facing this prenatal situation will likely proceed differently than he or she would in the postnatal period. The obstetrician will use array-CGH instead of FISH in order to cast a wider net—one that can catch a deletion on chromosome 22, as well as other possible deletions which may cause the heart defect.

Right now, the available array-CGH platforms can detect more than 40 syndromic chromosomal disorders. Just as with FISH, a normal result rules out only those conditions that correspond to the deletions or duplications that are covered on the array.

How Array-CGH Works

The technique involves labeling the patient's DNA in one fluorescent dye, labeling DNA from a normal control with a different fluorescent dye, allowing the DNA from both to mix, and then applying the mixture to a slide that contains small segments of DNA from known chromosomal regions.

The slide serves as the platform or the array. The mixture of the patient's DNA and the normal control DNA is allowed to match up, or hybridize, with the complementary DNA segments on the slide.

A scanner then reads the intensities of the two different dyes, determining their relative strength at each of the DNA spots on the array. If a patient has less DNA in a specified region of the genome—a deletion of chromosomal material—then the color of the control sample will be stronger at that point on the array. If a patient has more DNA in this specific region—a duplication of chromosomal material—then the color of the patient's sample will be stronger at that location.

Analysis can be performed on direct chorionic villi or amniotic fluid, or alternatively on cultured cells. For direct analysis, it might be necessary to amplify the amount of DNA obtained before running it on an array. In this case, it is essential that the amplification is uniform and does not introduce any bias.

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