Pre-implantation Genetic Diagnosis (PGD), also known as Embryo Screening, is a procedure used in conjunction with in vitro fertilization (IVF). PGD is generally recommended to detect numerical or structural anomalies in the chromosomes of embryos, as well as certain conditions caused by single gene defects. When embryos are affected by chromosomal conditions, these can prevent implantation to the uterine lining, lead to pregnancy loss, or result in the birth of a child with physical problems and/or mental retardation. PGD can help prevent adverse outcomes by identifying affected embryos as they are developing in the laboratory and before they are transferred to the womb during the IVF cycle. Our center offers PGD only after extensive consultation and review of your own clinical circumstances.

While there are multiple reasons for the failure of an embryo from IVF or natural conception to result in a viable pregnancy, the single most important factor is an abnormality of the chromosomes. Chromosomally abnormal embryos usually fail to implant in the uterus, while others implant but cannot develop normally and the pregnancy miscarries as early as in a few days to a few months. The timing of the miscarriage depends on which or how many chromosomes are abnormal in the embryo.

The percentage of chromosomally abnormal embryos that a couple produces varies depending on their own clinical circumstances. Factors such as advancing maternal age, the number of prior miscarriages, the number of failed IVF cycles, and the quality of sperm can directly influence the proportion of embryos that are abnormal.

Two different genetics techniques can be used during PGD to obtain diagnostic information necessary in deciding which embryos to transfer.

1. FISH (Fluorescence In-Situ Hybridization): A technique for "painting" whole chromosomes with a genetic probe bound to a fluorescent colored marker. FISH allows us to count the chromosomes based on color staining but it is limited to staining only 8-12 of the 23 chromosomes in the cell.
2. PCR (polymerase Chain Reaction): Is a technique whereby the chromosomes are split into pieces allowing for identification of normal and abnormal chromosome fragments. PCR works very well with single gene disorders but does not help identify the number of chromosomes present.

Pre-implantation Genetic Diagnosis (PGD) for Chromosome Anomalies (Aneuploidy)

Chromosomes are the physical structures that contain the DNA and genes necessary for our development; they are located in an the area of the cell called the nucleus. Normal human cells including the cells form an embryo contain 46 chromosomes in 23 pairs. We receive 23 chromosomes from each parent (or from the sperm and the egg). The first 22 pairs of chromosomes are the same for men and women. The 23rd pair determines our sex. A female has two "X" chromosomes, whereas a male has an "X" and a "Y." As such, the woman can only pass an X to her child in her egg. The man passes either the X or the Y in the sperm, therefore determining the sex of the child. Our reproductive cells (sperm and eggs) form by dividing into two cells through a process called meiosis. Each sperm and egg contains ½ of the chromosomes. In a normal conception, the egg and sperm contribute exactly 23 individual chromosomes each; one of each of the 22 numbered pairs and one from the sex chromosome pair. When an egg with 23 chromosomes fuses with a sperm with 23 chromosomes, the correct chromosome number of 46 (23 pairs) is again present, and the fertilized embryo has the best possible chance of developing appropriately.

If an error occurs, and the egg or sperm has an extra or missing chromosome, the embryo created by that egg or sperm will have an extra or a missing chromosome resulting in a condition called Aneuploidy. Sometimes similar errors occur despite the fact that the sperm and the egg have a normal chromosome count.

Having an extra chromosome is known as trisomy (tri=three instead of the normal pair) of a given chromosome, and having a chromosome missing is known as monosomy (mono= only one of the two chromosomes). When aneuploidy involves the larger chromosomes, the embryo may not attach to the wall of the uterus or it may stop developing soon after attaching and miscarry. However, if the aneuploidy involves chromosomes such as the 13, 18, 21, X or Y, the pregnancy may still carry on until birth, even though the pregnancy has a chromosomal disorder.

The most common of these is an extra number 21, known as Down syndrome or trisomy 21 (three 21 chromosomes). Other common aneuploidies are Klinefelter syndrome (XXY), trisomy 13, and trisomy 18. The features of the chromosome condition depend upon which chromosome is extra or missing, but can include physical differences and mental retardation.

Aneuploidy and Advanced Maternal Age

As a woman advances in age, the chance of aneuploidy in her pregnancies increases because her eggs are as old as she is. Females have all of their eggs in the fetal stage, and as a result, they are born with all the eggs they will have in their lifetime. In males, sperm is made every 65-75 days; therefore, the sperm is not as old as the man. Based on this, the theory regarding aneuploidy risk and advancing maternal age is that, over time, the chromosomes in the egg are less likely to divide properly, which results in the egg having an extra or missing chromosome. Therefore, the risk of aneuploidy increases with maternal age.

Aneuploidy Rate for 9 chromosomes (X, Y, 13, 15, 16, 17, 18, 21, 22)
in IVF-derived embryos (Data for 3600 cases)



As a woman ages, her fertility potential decreases and she also becomes more likely to miscarry when a pregnancy does occur. The process of cell division for reproductive cells (eggs and sperm) is called Meiosis, after meiosis, a "daughter" cells contains half the number of chromosomes as the parent cell. Faulty meiosis or non-disjunction occurs more commonly in eggs than sperm and results cells with the incorrect number of chromosomes. Meiotic nondisjunction increases in frequency as a woman ages such that as many as 60-80% of pregnancies miscarry when a woman is older than 45.

The purpose of preimplantation genetic diagnosis for aneuploidy, therefore, is to select for transfer only chromosomally normal embryos so as to achieve more pregnancies, reduce the number of miscarriages, and reduce the number of affected offspring.

PGD, using the FISH technique to count selected chromosomes can identify chromosomally-abnormal embryos. By selecting embryos containing the normal number of chromosomes (for those chromosomes tested), we can reduce the incidence of miscarriages and the incidence of babies born with abnormal numbers of chromosomes like trisomies (for example, Down's syndrome) and monosomies (for example, Turners syndrome). In certain cases where the woman produces a large number of embryos, we might also increase the pregnancy rate from IVF by increasing the probability that the transferred embryos are chromosomally normal for those chromosomes tested. THE PROCESS OF PGD

Two procedures are currently available in order to do PGD of embryos. Our PGD team which includes doctors, geneticists and embryologists will help you decide which procedure to use depending on your own clinical circumstances.

Polar Body Biopsy

The maturing egg produces two small cells, called the polar bodies, which degenerate after fertilization. The chromosomal content of these cells allows us to infer the chromosomal content of the egg. In order to test the polar body, an opening is made in the covering of the egg with the help of a precise microscopic laser and the polar body is carefully removed; all these procedures are performed by micromanipulation. The polar body is then analyzed while the egg is kept in culture in an incubator. Analysis of polar bodies provides information only from the mother. Chromosome abnormalities that may occur after fertilization (when the sperm meets the egg) will not be detected via Polar body biopsy.

Day 3 Embryo Biopsy and Blastocyst Biopsy

To test an early (3 day-old) developing embryo, a blastomere or embryonic cell is removed via a microscopic opening made a laser in the covering of the embryo during its third day of development (5 to 8-cell stage). The embryo is then kept in culture in an incubator while the cell is analyzed by PGD. More advanced embryos (blastocysts) can also be tested with the advantage of performing the test in more than one cell (usually 3 to 5) improving it's accuracy.

The PGD Analysis For Chromosomes

The biopsied cells are analyzed using a technique called "fluorescence in-situ hybridization" also known as "FISH". This technique uses probes, small pieces of DNA that are a match for the chromosomes we want to analyze, to count the chromosomes present. Each probe is labeled with a different fluorescent dye. These fluorescent probes are applied to the biopsied cell and attach to the chromosomes. Under a fluorescent microscope, the number of chromosomes of each type (color) can then be counted. The cyto-geneticist, therefore, can distinguish normal cells from cells with aneuploidy. Testing of the cells destroys them because they must be glued to a glass slide and repeatedly heated and cooled. As such, one cannot use them for another purpose or return them to the embryo. This analysis causes no extra inconvenience to the patient as it is accomplished in one day. The picture displays various chromosomes during FISH analysis.

Advantages and Clinical Application of PGD for Aneuploidy

1. Reduction in the Chance of Having a Child with Aneuploidy: According to current figures, the chance for a woman delivering a baby with aneuploidy is on average 1% if she is 35-39 years of age and approximately 3.5% if she is 40-45. PGD decreases the chance of having an affected baby. However, PGD is not capable of testing all of the 23 chromosomes at present. We therefore recommend that prenatal testing be performed in the resultant pregnancy via chorionic villous sampling or amniocentesis in order to confirm the diagnosis from PGD and to rule out other aneuploidies for which PGD can not test.
2. Increased Implantation Rate: It is well known that the pregnancy rate after in vitro fertilization decreases dramatically with maternal age. Aneuploid embryos have much lower survival rates than normal embryos, and half of them (the ones missing a chromosome) seldom implant. It appears likely that the decrease in pregnancy rates with maternal age is mostly caused by a corresponding increase in the number of aneuploid embryos. By performing PGD for aneuploidy and transferring only chromosomally normal embryos, the pregnancy rate might increase markedly. In several recent studies, an increase in implantation rates after PGD has been demonstrated.
3. Reduction in Miscarriages: In women 35 and older, approximately 35% of pregnancies are miscarried. Aneuploidy accounts for 50%, or more of these losses. By transferring only chromosomally normal embryos, the number of pregnancies going to term should increase. Recent studies have detected a significant reduction in pregnancy losses after PGD, from 23% to 9%. The increase in implantation rate and the significant decrease in pregnancy loss rate resulted in a significant increase in ongoing pregnancies and delivered babies. Some women repeatedly loose pregnancies because they produce eggs with abnormal numbers of chromosomes. In approximately 70% of spontaneous miscarriages an abnormal number of chromosomes is identified. The most common chromosome abnormalities found in miscarriages include trisomy 16 (3 copies of chromosome number 16); trisomy for chromosomes 22, 21, 15, 18 or 13; triploidy (3 copies of all the chromosomes); and abnormalities of the sex chromosomes. Another group of women and men with a history of repeated pregnancy loss may carry a genetic abnormality called a translocation. An individual with a chromosomal translocation is normal but an examination of their chromosomes pieces of two different chromosomes might have swapped places. Thus, during meiosis when the chromosomes split in half and one chromatid goes into one "daughter" cell and the other goes into the other "daughter" cell; we end up with a sperm or egg in which the total chromosome complement could be normal, a balanced translocation like the parent, or an unbalanced translocation which will fail as a pregnancy. FISH probes for the tips of the chromosomes (telomeres) are used to identify embryo status for translocation cases.
4. Repeated IVF Failures: In some couples successful delivery of a baby remains elusive even with the most intensive and sophisticated treatments. Identification of chromosomally abnormal embryos from the pool of embryos for transfer may improve the probability of a successful IVF treatment cycle. The incidence of aneuploid (abnormal) embryos is as high as 50% and similar to patients with recurrent pregnancy loss.
5. Family Balancing: The sex of an embryo can be determined by PGD and FISH prior to replacement in the womb. Our center offers this option also known as "Family Balancing" for those families where the number of boys/girls is uneven and the family would like to choose the sex of their next child. In addition, combining PGD with MicroSort, an experimental technique to sort the sperm cells into the X & Y-bearing fractions, increases the number of embryos of the desired sex.

Identification of Genetic Conditions with PGD

Over 10,000 human diseases are caused by defects in single genes. Eventually, every genetic disease with an identified gene mutation will be detected by PGD. Single gene disorders are individually very rare and they affect a very small part of the general population as a whole. Since only a single gene is involved in each case, these conditions generally have simple inheritance patterns in family pedigrees. This means that they can be traced through families and their occurrence in later generations can be predicted. The defective version of the gene responsible for the disease is known as a mutant allele. Single gene disorders can be divided into a number of different categories according to how they are transmitted from generation to generation. Some are described as dominant diseases because only one mutant allele is required, and such diseases tend to turn up in every generation. Other diseases are described as recessive because both copies of the gene must be defective in order for the disease to occur. These recessive diseases often skip generations because the mutant alleles can be carried without any effect if a normal allele is also present. Many single gene disorders affect both sexes equally. However, where the relevant gene is present on the X-chromosome, the associated disease tends to be more common in males.

An individual with a family history of a single gene disorder may be at risk for passing the condition onto his or her children. Examples of single gene disorders include cystic fibrosis, sickle cell anemia, Tay Sachs disease, myotonic dystrophy, Duchenne and Becker muscular dystrophies, Fragile X syndrome and spinal muscular atrophy, among many others. An individual that carries an abnormal gene in their chromosomes does not necessarily have the disease because they only have one copy of the abnormal gene; these conditions are termed autosomal recessive and are the most common form. However, if they create a pregnancy with another person who also carries the same abnormal autosomal recessive gene then they have a 25% chance of having an affected child.

Other types of genetic disorders may sometimes occur (autosomal dominant, X-linked recessive, etc.). A consultation with a genetic counselor can help you understand the variations and risks for having an affected child. The genetic basis for some diseases such as cystic fibrosis, spinal muscular atrophy or Huntington's chorea resides in a single gene.

PGD for single gene conditions allows the identification of unaffected embryos prior to their placement into the womb, therefore, reducing the risk of having a child with the genetic disorder. The list of genetic conditions for which embryos can be tested is growing very rapidly due to the availability of new tests; basically as long as a specific gene mutation is identified for a particular condition, a specific probe that binds and detects that particular region on the DNA can be developed and tested prior to performing testing on the embryos. This phase is usually done using a couple of blood samples form the prospective parents way in advance of their actual IVF-PGD cycle.

There are many conditions for which single gene testing is already available; please contact us directly if you require additional information, or if a specific condition is not listed on our website yet. Our center offers PGD only after extensive consultation and review of your own clinical circumstances. At our facilities you will be in touch with a multidisciplinary team that includes genetic counselors, psychologists, physicians, embryologists and cytogeneticists which will help you prepare and undertake your IVF-PGD treatment cycle.

Conditions for which PGD is available

Risks and Limitations associated with PGD

1. The Risk of Embryo Biopsy: While PGD is a relatively new procedure in IVF, the micromanipulation techniques required to perform it have been in use for many years. The risk of accidental damage to an embryo during removal of the cell(s) in the hands of an experienced embryologist is very low, and it is currently calculated at less than 1.0%. Other Assisted Reproduction procedures such as Intracytoplasmic Sperm Injection (ICSI), Fragment Removal and Assisted Hatching are all performed by making microsurgical openings in the covering of the egg or embryo and none have been found to have other than mostly positive effects on implantation and viable pregnancy rates.
2. Removal of Cells from the Embryo: No part of the future fetus will be affected because one or two cells are removed from an embryo approximately two days after fertilization. At this developmental stage all cells in an embryo remain totipotent; that is, they have not differentiated yet, meaning that each cell by itself can grow into a whole and perfect fetus. The biopsy procedure merely delays continued cell division for a few hours, after which the embryo reaches the same number of cells as before and continues its normal development. It is possible that embryo biopsy may lower embryo implantation rates slightly, while selection of chromosomally normal embryos via PGD may increase them. Therefore, the balance between potential biopsy damage and beneficial effects of PGD seems to be positive.
3. Misdiagnosis: The accuracy of PGD for aneuploidy is approximately 90-95%. This means that the error rate is approximately 5-10%. Within this chance of misdiagnosis, there is a false negative rate, a false positive rate, the chance for no result and the chance for mosaicism. A mosaicism is defined as the embryo having cells with different chromosome make-up. Typically, all cells of the embryo have the same chromosomal make-up as they originate from the same fertilized egg. However, it is possible for cells of the same embryo to have differing numbers of chromosomes. When the cell analyzed has a different chromosomal complement than all the others in the embryo a misdiagnosis occurs. Due to the chance of misdiagnosis as well as the presence of aneuploidies, for which testing is not available, we recommend prenatal testing as stated earlier.
4. Technical and biological Aspects: There are certain technical and biological issues which may reduce the accuracy PGD. For example, the cell must contain an intact nucleus which contains the chromosomes. If the cell has no nucleus or if the nucleus ruptures during the biopsy procedure or cell fixation process, that cell cannot be evaluated. The incidence of non-diagnostic cells varies between 5-20%. If the cell chosen for biopsy happens to be in the middle of cell division, the result may show four sets of chromosomes and falsely lead to a diagnosis of an abnormal embryo. Sometimes, two chromosomes might be lying on top of each other; this may lead to an underestimate of the number of chromosomes and therefore a false interpretation. In some instances, the fluorescent probe does not bind to a chromosome suggesting a missing chromosome. Some embryos may have two separate cell lines. These embryos are called to be Mosaic. Mosaic embryos may be abnormal but in many cases the abnormal cell line dies off or becomes part of the placenta rather than the baby. Unfortunately, we cannot tell the final outcome. Consequently, a mosaic embryo could be read falsely as normal or abnormal. In total, we believe that the false normal rate is about 5% and that the false abnormal rate is about the same, 5%.