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Spanish Researchers Develop 'Short CGH' Approach for Preimplantation Genetic Screening


By Justin Petrone

Scientists at the Universitat Autònoma de Barcelona in Spain have developed a new comparative genomic hybridization approach for screening embryos prior to implantation during an in vitro fertilization cycle.

The technique, called short CGH, has been applied to the screening of chromosomal anomalies in cases of advanced maternal age, recurrent miscarriages, or repeated implantation failures.

The authors have described short CGH in two papers, most recently in Fertility and Sterility in January. A third paper is in press.

The method relies on metaphase CGH slides sold by Abbott Molecular, and could be seen as an alternative to array-based approaches currently gaining acceptance for PGS (see related story, this issue). The authors claim the approach is faster than traditional techniques like fluorescent in situ hybridization, but cheaper than array-CGH.

FISH is still the most commonly used methodology for screening embryos for anomalies during IVF cycles. One cell of the embryo is typically analyzed on the third day of its development, though FISH only analyzes nine of the 24 chromosomes. The implantation rate using this technique is 14 percent.

CGH offers an alternative because it can assess all chromosomes, but it requires 72 hours to yield results, meaning that embryos analyzed must be frozen and transferred in a subsequent IVF cycle, which could affect their viability. The implantation rate using this technique ranges from 40 percent to 70 percent.

UAB's new short CGH approach enables the analysis of the entire karyotype in 12 hours, allowing embryos to be selected and transferred within the same IVF cycle.

"We wanted to perform a comprehensive analysis of all chromosomes from the human karyotype in the PGS but avoid the cryopreservation step required when standard CGH is used," Mariona Rius, a researcher in UAB's Cellular Biology and Medical Genetics Unit, told BioArray News recently.

"With the short-CGH approach we achieve the full karyotype analysis within approximately 12 hours, so fresh embryo transfer is possible in the same IVF cycle," she said.

Rius said she and fellow researchers have been using traditional CGH for PGS since 2004. "We were skilled in performing standard CGH and, therefore, the best way was adapting our protocol to achieve a reduction of the hybridization period from 72 hours to 12 hours," she said.

Rius added that short-CGH is a "reliable technique to be applied in PGS" and touted the approach as being "cheaper than array-CGH," though she did not elaborate.

The UAB researchers use Vysis CGH metaphase slides from Abbott Molecular. The technique is accomplished by hybridizing control and test DNA to normal metaphases that are fixed on the slide, capturing the hybridized metaphases, karyotyping them, and evaluating the ratio between green and red fluorescence, Rius said.

The process is accelerated by a 30-minute exposure in a microwave, and treatment in a moist chamber in rotary agitation for 12 hours. A minimum of 12 metaphases per cell-case are subsequently captured and analyzed using Abbott's Isis CGH software.

The approach was validated using fibroblasts isolated from confluent cultures of cell lines with known aneuploidies, Rius said. Afterwards, short-CGH was validated using discarded embryos from FISH-PGS cases.

Once the protocol was validated it was applied in PGS clinical cases. Two children have been born following PGS performed by short CGH and there are two ongoing pregnancies, Rius noted.

Rius said that she is confident that the short-CGH method will be applied in more PGS tests. "UAB does not work as a PGD center, but in our group we are currently applying this strategy in several cases that contribute to enlarge the scientific progress," she said.

"This method is now accessible to everyone who wants to carry out PGS. But some IVF clinics in Barcelona are currently working with our group as part of our scientific project and they can benefit from our short-CGH PGS," she said.

Nick Haan, CEO of Cambridge, UK-based BlueGnome, which currently sells arrays and array-based services for PGS, acknowledged that the UAB researchers have created a "highly developed" metaphase protocol that "significantly reduces" the timescale of mCGH.

Still, Haan told BioArray News that while short-CGH is a "useful advance, arrays retain advantages in that they are less labor intensive, less subjective, and can be more sensitive."

While arrays are "more costly in terms of consumables than mCGH, the difference is minor and labor costs are much reduced since arrays are amenable to automation," he said.

Have topics you'd like to see covered in BioArray News? Contact the editor at jpetrone [at] genomeweb [.] com.