Preimplantation genetic testing (PGT)

Preimplantation genetic testing (PGT)

PGT is a very early genetic diagnosis method to select healthy IVF embryos before transferring them into the uterus. Nadezhda hospital’s Genetics lab is equipped with the necessary technology and expertise to guarantee GMP compliance and allow genetic testing in-house.


The team of our Genetics lab was the first to introduce PGT in Bulgaria back in 2008. Since then, we have successfully analysed thousands of embryos and have achieved a solid experience in this type of test.


PGT is recommended in mutations that result in miscarriages, stillbirth, or life-threatening conditions and diseases with severe physical or mental impairments, for which currently there is no treatment.

PGT for monogenic disorders (PGT-M)

This type of preimplantation testing is indicated in families where one or both partners are carriers of monogenic defects for diseases with different inheritance types: autosomal recessive, autosomal dominant, X-linked (sex-linked), and hereditary cancer syndromes (germline mutations). The risk for the birth of affected offspring is high with each pregnancy and varies between 25 and 100%.

With two parents who are both carriers of an autosomal recessive mutation, the risk for transmission to the offspring is 25%. Diseases with this inheritance type are beta-thalassemia (Cooley’s anaemia), cystic fibrosis, epidermolysis bullosa, spinal muscular atrophy, various metabolic disorders, etc.

With this type of inheritance, the risk is 50% for every child. This group includes Charcot-Marie-Tooth disease, neurofibromatosis, myotonic dystrophy, Huntington’s disease, osteogenesis imperfecta, familial adenomatous polyposis, and achondroplasia, among others.

The risk can vary between 25% to 50% depending on the inheritance type, recessive or dominant. Diseases in this group include Duchenne muscular dystrophy, fragile X syndrome, haemophilia, incontinentia pigmenti, focal dermal hypoplasia, or Goltz syndrome. These disorders only affect one of the sexes, so healthy embryos of the opposite sex could be selected for transfer. Another option for selecting healthy embryos is DNA analysis for the specific mutation if available technology would allow it.

PGT for structural rearrangements (PGT-SR)

PGT-SR is recommended for couples in which one of the partners has a chromosomal abnormality (a balanced structural translocation, inversion or other chromosomal rearrangements; mosaicism in one of the partners). The risk for the resulting embryos is high and varies according to mutation type.


In about 2–4% of couples with recurrent miscarriages, one of the partners is a carrier of the so-called balanced chromosome mutations – reciprocal and Robertsonian translocations, inversions, etc. This type of mutation is called familial because it is usually inherited by a healthy parent and is seldomly new. The carrier is typically healthy, but a certain percentage of their sex cells have a genomic imbalance. Such gametes have a poor fertilising capacity, resulting in implantation failure, miscarriages or the birth of offspring with chromosomal disorders. With carriers of chromosomal rearrangements, the miscarriage risk is higher with every subsequent pregnancy, varying according to the mutation type.


Since all the embryo’s chromosomes are usually tested, not only those involving the chromosomal rearrangement, the technique provides an opportunity to discover additional mutations in the remaining chromosomes.

PGT for aneuploidies (PGT-A)

 Indications for PGT-А include advanced reproductive age of the female, recurrent miscarriages, previous failed IVF cycles, severe male infertility.

In PGT-А, embryos are analysed for the total number of chromosomes. Approximately 70% of embryos resulting from spontaneous conception or an IVF procedure are lost before birth. Most of them are lost during the first trimester, some even before implantation. The primary cause for this is chromosome abnormality known as aneuploidy, resulting from the loss or addition of chromosomes.

At least 85% of aneuploidies originate in the egg. Sperm have a significantly lower contribution, to only 7-8% of all cases. The remaining aberrations occur accidentally during cell division in the early embryo stages.

It is known that, on average, more than 50% of embryos have a chromosomal abnormality. Since this percentage increases with age, this is probably the main reason women have difficulties conceiving and have more miscarriages with advancing age. The high incidence of chromosomal mutations adds to the benefit of doing PGT-A of embryos during IVF.

Some chromosomal abnormalities allow the pregnancy to be carried to term and live birth to occur (Down syndrome, Edwards syndrome, Turner syndrome, etc.); others result in preterm birth, miscarriage or embryonic development arrest even before implantation.

The high frequency of chromosome mutations makes PGT-A a vital tool in the course of IVF treatment.

Stages of the PGT procedure

PGT can be performed only with in vitro fertilisation (IVF). The IVF for PGT itself includes controlled ovarian stimulation, oocyte retrieval by follicular aspiration, collection of sperm from the male partner, egg fertilisation, embryo biopsy, PGT analysis and embryo cryopreservation.

Different biopsy techniques are used, such as polar body biopsy, blastomere biopsy of early embryos, and trophectoderm biopsy of blastocyst embryos

Embryo biopsy is done on day 5 or 6 after fertilisation, at the blastocyst stage. Five to seven cells are biopsied of the outer blastocyst layer, the predecessor of the placenta and fetal membranes. In this way, the embryo is kept intact and frozen afterwards. If necessary, frozen embryos can be biopsied too after thawing. When an experienced embryologist performs the procedure, the risk for embryo arrest is less than 1%.

What follows is DNA isolation from the biopsied cells, amplification and analysis by a suitable genetic test depending on the mutation of interest. Results typically take two to four weeks, and their precision is over 95%. Free pre- and post-test genetic counselling is available to all patients of Nadezhda Hospital.

Different biopsy techniques are used, such as polar body biopsy, blastomere biopsy of early embryos, and trophectoderm biopsy of blastocyst embryos.

The biopsy is done using a laser to make a tiny opening into the egg’s outer layer, through which one or two polar bodies are aspirated. This approach is informative only about maternal mutations, as it does not account for the paternal genome contribution or for mutations arising after fertilisation. The procedure typically does not affect the further development of the embryos.

Typically only one or two cells are extracted by the biopsy on day three after fertilisation when the embryo is at the 6-8 blastomeres stage. The main disadvantage of this approach is the relatively high incidence of natural mosaicism in 3-day embryos. It is possible that the analysed cell has defects while all the rest are normal; as well as the opposite scenario – the analysed blastomere to be normal, while the remaining cells have defects. In PGT-M, the isolation and amplification of DNA from a single cell also bear risks of ADO (allelic dropout – a partial amplification failure affecting one allele). This would result in misdiagnosis and is the main reason why this approach has been losing popularity in clinical applications.

This is the most popular approach currently. It is performed on day five or six after fertilisation, at the blastocyst stage when the embryo consists of 100–150 cells. The blastocyst is like a balloon with a small ball inside, composed of two layers of cells. The cell group on the inside is the one from which the foetus develops. The outer surface cells surrounding the cavity are called the trophectoderm, and they give rise to the future placenta. In trophectoderm biopsy, 5–7 cells predecessors of the future placenta are taken, and no future foetal cells are affected. The advantage of trophectoderm biopsy is that the analysis uses a larger number of cells and has higher precision and a lower risk of misdiagnosis due to mosaicism and ADO.

Preimplantation genetic testing bears minimal risks for the further development of the eggs and embryos. The risk of biopsy-related damage is less than 0,2% in oocytes and less than 1% in embryos. Approximately 5% risk of misdiagnosis exists (a false positive or a false negative) due to the embryo’s natural mosaicism or technical limitations of the method used. The analysis results could indicate that there is not one normal embryo of all tested so no embryo transfer can be done. In some cases, results could be impossible to interpret.


PGT success rates directly correlate with the success rates of the IVF clinic; therefore, PGT should be attempted only ат the best IVF clinics after in-depth medical genetics counselling. If pregnancy occurs, in all cases, non-invasive prenatal testing is recommended to confirm PGT results. There is no increased risk of congenital anomalies in children born from a PGT-IVF procedure.

Currently, PGT of polar bodies, blastomeres and trophectoderm cells is available at Nadezhda hospital. Different methods can be used depending on the genetic mutation that needs to be analysed. The genetics lab has equipment available for all current technologies of genetics analysis:

  • Fluorescent in situ hybridisation (FISH);
  • DNA microchip analysis (Array comparative genomic hybridisation (aCGH));
  • DNA Real-time polymerase chain reaction (RT-PCR);
  • Next-generation sequencing (NGS).

See also

Genetic tests for infertility
Prenatal genetic diagnosis
Other diagnostic and screening tests
Heterozygote carrier screening tests