Genetic research of the embryo during IVF. Preimplantation genetic diagnosis (PGD) For IVF. Stages of PGD

Many potential parents planning in vitro fertilization are interested in the question: PGD in IVF - what is it? What is the essence of this procedure and is it as important and safe as doctors say?

The abbreviation PGD encodes the name of embryonic research - preimplantation genetic diagnosis. Thanks to this procedure, you can confidently guarantee the successful implementation of IVF. During the PGD analysis, each embryo is carefully checked for genetic abnormalities.

But is this research really necessary in the IVF cycle? Maybe you can do without PGD? The choice is always with the parents-to-be, but it should be remembered that caution is never excessive. Especially in such a matter as conception in the laboratory. It should not be assumed that if the embryo cultivation process was successful, then the successful outcome of the entire IVF procedure is inevitable. There is no guarantee that all embryos obtained are free of genetic abnormalities and have sufficient vitality. The transfer of defective embryos can provoke:

• developmental deviations in children due to IVF;
• inability to implant the embryo;
• spontaneous termination of pregnancy;
• freezing of the fetus.

PGD ​​significantly increases the chance of having an absolutely healthy baby as a result of IVF.

Pros of PGD

The main advantage of PGD is the ability to analyze the state of health of the embryo intended for IVF prior to implantation into the uterine cavity. Preimplantation genetic diagnostics allows:

• to double the percentage of successful implantations during IVF;
• to minimize the chance of occurrence of genetic deviations;
• reduce the likelihood of termination of pregnancy by 25%;
• select the most viable embryo for IVF;
• weed out embryos with genetic abnormalities;
• determine the reasons for the failure of the previous IVF;
• determine the Rh factor;
• prevent an unplanned multiple pregnancy;
• identify the presence of extra or missing chromosomes;
• to establish the sex of the unborn child, in order to prevent the transmission of sex-related diseases during IVF.

Indications for PGD in IVF

Couples with a high risk of transmitting genetic abnormalities become candidates for the passage of PGD embryos. IVF diagnostics is necessary if:

• at least one of the two parents has genetic diseases;
• the mother is over 40 years old;
• father is over 45 years old - with age, the number of sperm with deviations increases;
• there have already been unsuccessful IVF attempts in the past;
• spermatogenesis problems in men;
• IVF ICSI fertilization;
• death of the implanted embryo during previous pregnancies due to the mismatch of the Rh factor.

Methods for preimplantation genetic diagnosis of embryos

Various types of PGD are practiced during IVF. Each of them pursues specific goals and has fundamental differences.

PGD ​​options

FISH is a method for molecular genetic analysis. This method allows you to exclude unbalanced chromosomal abnormalities of the embryo intended for IVF - an altered number of chromosomes (aneuplody) and structural changes (translocations). The blastomere extracted during the biopsy is placed on a glass and heated to the desired temperature. Then it gradually cools down. As a result, the integrity of the shell is violated. PGD ​​is characterized by the use of FISH probes. These are a kind of fluorescent labels that mark sections of DNA. Then, using a special microscope, the number of chromosomes is determined and abnormally developing structures are identified.

NGS is a revolutionary diagnostic method that allows you to examine 23 pairs of chromosomes at once. Due to the complexity of this type of PGD, the price for the service is quite high. The errors of this method are insignificant - within 0.1%. Diagnosis takes place in an IVF cryocycle without biopsy. Allows you to accurately establish the presence of genetic pathology by examining the DNA sequence. In the process, existing mutations, monogenic anomalies, and translocations are identified. This is the most expensive method of PGD in IVF.

PCR is a polymerase chain reaction technique. It is the leading PGD method for detecting monogenic abnormalities. DNA is denatured to unwind the double strand. Then, a fragment of a nucleotide is removed and, using specific enzymes, the amount of the starting material is doubled and defects in the structure of chromosomes are detected. PGD ​​is indicated in cases where one or both potential IVF participants act as carriers of diseased genes, in order to exclude their transmission to the child.

CGH is a comparative genomic hybridization technique. One of the most efficient and costly PGD options. In the process of research, similar DNA strands are hybridized into a single molecule for comparison. The test and control DNA during PGD are labeled with different fluorescent dyes. Computer analysis during PGD reveals various defects - loss, duplication or transfer of a chromosome section (deletions, amplifications, translocations and other types of mutations).

The above PGD methods are not interchangeable and are used to identify different deviations. Only a doctor performing IVF can determine the relevance of a particular procedure for each specific case.

Stages of PGD

If during IVF there is a need for PGD, the couple receives advice from a specialist - a geneticist. Parents-to-be are tested for karyotype and mutations. Indications for PGD are revealed. The doctor then advises the couple on the key features of the IVF diagnostic procedure.

Diagnostic stages

At the first stage, the woman is stimulated with hormones and transvaginal egg retrieval - according to the standard IVF plan. Injection of a sperm cell into a female cell by the ICSI method. This technique is used because of the need to weed out spermatozoa that are not involved in the IVF process.

Biopsy for PGD with extraction of the blastomere and its subsequent fixation. On the third day, one of the formed 8 cells is cut off from the embryo. The procedure is performed using a mechanical, laser or chemical method. The cells during this period are not differentiated and are identical. Each of them has a complete set of genetic material. Therefore, the biopsy does not prevent the remaining group of cells from developing fully.

Conducting PGD using the methods shown. The study lasts no more than 2 days. On the 5th day, the embryo can no longer remain in the incubator. It is necessary to analyze and present the results of PGD as soon as possible in order to carry out the transfer of the embryo into the mother's uterus.

PGD ​​results

Based on the results of the diagnostics, embryonic units with pathological changes in chromosomes are eliminated. The result directly depends on the age of the expectant mother. With PGD of embryos, women who are in the reproductive period - up to 40 years old, approximately 15-20% are discarded. At the age of over 40, with a past experience of unsuccessful IVF and gene mutations in the parents themselves, 100% may be unsuitable for implantation.

Cost of PGD embryos service

PGD ​​is an expensive procedure, but for certain indications it is simply necessary for the successful completion of an IVF cycle. It eliminates the risk of unsuccessful replanting, reproduction of sick offspring. In fact, it saves money, which, in case of failure, might be needed for a second IVF procedure, and in the case of a baby with a genetic deviation, for treatment and care.

The cost of PGD in Moscow at the "IVF Center" varies depending on the research method and the number of embryos diagnosed. The significant cost of PGD is due to the increased complexity of the procedure and the use of expensive equipment, drugs and consumables.

If there is a history of unsuccessful replants, or the couple has already had children with genetic abnormalities, PGD is simply necessary. To perform IVF with PGD in Moscow, contact our clinic by phone or sign up for a consultation with an IVF specialist through a special form on our website.

Start your journey to happiness - right now!

Currently, there is a rapid development in all areas of various innovative technologies, technical and scientific progress has not passed by medicine. In the field of reproductive medicine, doctors have achieved tremendous success. Nowadays, various methods of assisted reproductive technologies are common, although earlier humanity could not even dream of such a thing. One of the types of such technologies is the use of the in vitro fertilization method. This gives hope to all infertile couples who have already tried all the treatments and methods of conceiving, but which were only a waste of time. The in vitro fertilization protocol has a number of rather complex drug regimens and manipulations, is a laborious process, and is also accompanied by significant psycho-emotional stress for the woman and the family as a whole. Various congenital malformations of the fetus, chromosomal abnormalities are not so uncommon for doctors now. Which are detected at the first, at the second screenings, and some can be identified after the birth of the child, even in the case of a spontaneous pregnancy. Therefore, one of the main tasks in modern reproductive medicine is the development of the industry for identifying various pathologies of the development of the embryo even before the stage of its transfer into the cavity of the mother's uterus. For this purpose, fertility doctors use the so-called IVF with PGD. IVF with PGD in Moscow is available in the same way as in other large cities of the Russian Federation.

Genetic analysis of the PGD IVF embryo

What is it and what is the essence of the technique?

PGD ​​stands for preimplantation genetic diagnosis - genetic analysis of an embryo during IVF at the stage of its cultivation outside the mother's body. With the help of this study, embryologists select embryos with unsatisfactory performance and are excluded as material for transfer. Further work continues with high-quality embryos without any genetically abnormalities. The risk of developing a child with any genetic abnormality is minimized.

However, not everyone and not always are offered a pre-implantation study. A group of factors is distinguished, in the presence of which, the IVF protocol implies such a study.

These include:

1. If, entering into the protocol of in vitro fertilization, a woman has reached the age limit at 35 years old, then the risks of developing genetic abnormalities increase significantly, unlike women younger than this age;
2. For the same reason, embryos that were obtained by fertilization with spermatozoa from a man over 45 years old are subject to research. Risks increase every year;
3. A history of fetal death due to rezuz-conflict pregnancy;
4. Complicated hereditary history of at least one spouse from a couple. Since there is a high risk of hereditary transmission of genes encoding a genetic pathological condition.
5. Reproductive history of several in vitro fertilization flight protocols;
6. Complicated obstetric history in the form of spontaneous abortion. Dead pregnancies. This is due to the fact that such pathological conditions of pregnancy up to 8 weeks are due in 80% to genetic and chromosomal abnormalities;
7. In male sperm, the content of abnormal sex cells - spermatozoa - is increased.

This diagnosis allows you to identify a fairly large number of nosological units, such as achondroplasia, cystic fibrosis, Down syndrome, Edwards, Patau and many other anomalies.

Diagnosis requires a significant number of follicles. Therefore, with PGD, a woman is given increased ovarian stimulation.

What do fertility doctors want to achieve using such a complex and time-consuming technique:

These include biochemical and ultrasound screening, which calculate the risks of having a child with developmental pathologies. When high rates of prenatal screening are detected. The woman should be sent for invasive diagnostic methods, such as amniocentesis - amniotic fluid collection in order to obtain material for the study of the karyotype, cordocentesis - the study of umbilical cord blood.

Pros and cons of IVF PGD

Prenatal genetic diagnosis is positive. So are the negative sides.

The positive ones include:

1. Conducting a genetic study and transferring it to the uterine cavity with 95% confidence when carrying out the protocol of in vitro fertilization of healthy, viable embryos.
2. The possibility of obtaining offspring of the sex that the parents want.
3. The positive is the ability to prevent Rh-conflicts, as well as hemolytic complications of these pathological conditions.
4. Prevent spontaneous abortion due to chromosomal, genetic abnormalities, which in 80% are interrupted before 8 weeks.
5. Due to the absence of complications associated with various genetic abnormalities, the in vitro fertilization program ends up in large cases with success.
6. Congenital defects are almost completely excluded.
7. Another, quite an important point, which is greatly influenced by the receipt of preimplantation diagnostic data, is the woman's psycho-emotional peace. A poor mother, even one who got a spontaneous pregnancy, due to the availability of a variety of information, worries about the birth of a child with any developmental defect. These pathological conditions are more and more common every year among children born for a variety of reasons. Therefore, the mother, who received an answer that by 95% guarantees her the birth of a healthy child, will feel calm, confident and her psychological attitude will not affect the occurrence of uterine tone and, accordingly, reduce the risk of spontaneous abortions, threats of termination of pregnancy, and the threat of premature birth.

Like any method, like a medal, there are two sides, both there are good points and negative ones:

1. Absolutely negligible, but the risk of damage to the embryo remains, it is equal to 1%. But 99% of success with this procedure fully justifies them.
2. Nevertheless, despite the high information content of the method, a small percentage remains - about 5 errors. These missed points are corrected during screening studies.
3. A significant disadvantage of preimplantation genetic diagnostics is its price. The cost of PGD for IVF is about 60 thousand rubles and more, depending on the clinic and the diagnostic method.

In some types of PGD with IVF, the price reaches 140 thousand rubles. The high cost of this method depends on very expensive equipment, as well as reagents for conducting analyzes, which are used to identify pathologies of fetal development. In IVF with PGD, the cost also depends on the number of embryos studied, as well as the number of nosological units that will be examined.

However, this cost can never be compared to the cost of a human life. And especially the life of a still-born child.

IVF techniques with PGD - cost

Distinguish between different methods of carrying out pre-implantation genetic research. These include:

• The research method, coded by the abbreviation "FISH", is a fluorescent hybridization method that involves the study of a molecular genetic species. All genetic material is found in the nuclei of every cell. It is advisable to carry out diagnostics at certain periods of cell division, namely in the interphase or metaphase. This is the most affordable method from a financial point of view, however, its information content is the least, since it does not provide an opportunity to study the entire chromosome set, which is a rather negative feature of the method.
• Method. Which makes it possible to explore the entire chromosome set is CGH - a method of comparative genomic hybridization. It allows the identification of blastocysts with high implantation potential. But he also has a huge drawback - this is its price. The method is expensive and not affordable for all married couples entering the in vitro fertilization protocol.
• The PCR method - polymerase chain reaction, also diagnoses hereditary diseases as a result of mutations.
• NGS method - Now-Generation Sequencing, an innovative method that will soon replace all other methods. It is highly informative and reproductive specialists have high hopes for it.

What does a woman have to go through when conducting a genetic study? What are the stages of the method of this diagnostic manipulation?

First, a married couple should be consulted by a geneticist for a full examination, to identify indications for preimplantation diagnosis. Further, the protocol of in vitro fertilization takes place without any features that would affect women.

In vitro fertilization is carried out according to plan until the moment the egg and sperm are combined and the embryos are cultured. Three days later, the stage of pre-implantation diagnosis begins. Under special conditions, using microinstrumentation, embryologists obtain cells from cultured embryos. The method is safe and, in most cases, does not cause any complications.

Then, with the selected material, research is carried out by any of the above methods. This stage is one of the most responsible, since the further fate of pregnancy, the still-born child, mother, as well as all families depends on the qualifications of the reproductive doctor, on his choice.

After receiving the results, embryos that do not meet the requirements are screened out, and only high-quality, viable embryos are transferred. The replanting is performed on the 5th day of cultivation.

Preimplantation genetic diagnostics does not have any complications and the incidence of unsuccessful in vitro fertilization protocol without OGT does not differ from IVF with PGD.

This method is an excellent way to reduce the frequency of bearing children with congenital anomalies, however, for certain reasons, it is not always possible to carry out it due to the presence of contraindications to its use.

1. The embryo under study does not have six blastomeres, which is insufficient for PGD.
2. The embryo is diagnosed with multinucleated blastomeres;

IVF genetic diagnosis - what is it for

In general, the method is an excellent prevention of congenital malformations of the fetus, increases the chances of a successful pregnancy and further gestation without the threat of interruption, even prevents the development of res-conflicts, which is of great importance for Rh-negative women, for whom this protocol of in vitro fertilization is not the first or have already had a history of pregnancy. The only, but very significant, disadvantage is the price of the procedure. Not every infertile married couple can afford even the very procedure of in vitro fertilization due to financial difficulties, and even more so, additional expenses on preimplantation diagnostics. But at the moment there is a solution to this issue. According to the Federal Program, by submitting an application to the website, married couples can take part in the program of free IVF protocol with PGD under compulsory medical insurance, that is, IVF with PGD, the cost of payment for these manipulations is borne by the compulsory health insurance fund. This program gives a chance for a bright future to desperate families who have such a terrible diagnosis of infertility. Do not despair, because the greatest happiness has now become so close to you.

Preimplantation genetic diagnosis (PGD)

is an analysis of genetic disorders in embryos before their implantation into the uterine cavity... This is a new direction in the field of genetic diagnostics and reproductive technologies. The study is carried out at the earliest stage of human development, when the embryo is only a few days old and consists of several cells.

Preimplantation diagnosis of the embryo is carried out in the cycle of in vitro fertilization before embryo transfer into the uterine cavity... This prevents the transfer of genetically defective embryos. The main advantage of PGD is that it is performed even before pregnancy.

For the first time, preimplantation genetic diagnosis (PGD) was carried out in 1990. Currently, PGD is carried out in many countries that are actively developing assisted reproductive technologies.

PGD ​​can be performed in situations where there is an increased risk that the embryos will have certain chromosomal abnormalities. Such pathologies can reduce the likelihood of implantation (attachment) of the embryo in the uterus, lead to miscarriage, pregnancy fading or the birth of a child with physical or mental disabilities. PGD ​​helps prevent this adverse outcome by allowing healthy embryos to be selected prior to transfer to the uterus.

Many patients want to pre-select the gender of the child, which can also be done with the help of PGD.

Depending on the purpose and research methods, there are three main directions of PGD:

Diagnostics of numerical chromosome abnormalities (aneuploidy);
PGD ​​for translocations;
PGD ​​of monogenic defects.

In what cases is PGD carried out

Women over 35;
men over 39 years old;
married couples after repeated unsuccessful IVF attempts;
men with severe spermatogenesis disorders (single spermatozoa, high percentage of abnormal spermatozoa, etc.);
women with a history of recurrent miscarriage (especially in cases of early termination of pregnancy);
history of unsuccessful IVF attempts (more than two);
couples with a high genetic risk (family cases of hereditary diseases, karyotype abnormalities).

Diagnosis of numerical chromosome abnormalities (aneuploidy)

Diagnosis of numerical chromosome abnormalities (screening for aneuploidy) allows you to identify embryos with the presence or absence of one or more chromosomes.

Normally, all human genetic material is distributed in 46 chromosomes (23 pairs). At conception, the embryo receives half of the genetic material (chromosome 23) from the mother (the chromosome of the egg), and the other half from the father (the chromosome of the sperm). Abnormalities in the number of chromosomes in the germ cells of parents occur quite often and the future the embryo may receive an extra chromosome (this is called trisomy) or might happen loss of any chromosome (monosomy). Such violations are called aneuploidy. These defects lead to abnormalities in the development of the embryo at any stage: lack of implantation, spontaneous abortion, or the birth of a child with a severe hereditary disease, such as Down's disease (trisomy on chromosome 21). PGD ​​is performed for 9 chromosomes in which aneuploidy is most common. These are chromosomes 13, 15, 16, 17, 18, 21, 22, X, Y. Currently, it is not possible to diagnose all 23 chromosomes.

The causes of chromosomal abnormalities in the formation of gametes (eggs and sperm) remain poorly understood. At the same time, the risk factors for such disorders are known. These include the woman's age. With the age of a woman planning motherhood, the risk of egg aneuploidy increases. and, accordingly, the risk of having a child with trisomy, in particular with Down's syndrome, increases. Therefore, for pregnant women over 35 years of age, prenatal (prenatal) diagnosis is recommended in order to study karyotype (number and structure of chromosomes) of the fetus... If the fetus has a chromosomal abnormality, termination of the pregnancy is recommended, since the chromosomal defect is virtually untreatable and the child born will suffer from a serious illness with multiple birth defects in the absence of effective treatment.

The IVF procedure is often used by married couples in which the woman is over 35 years old. Therefore, prenatal or preimplantation diagnosis is recommended for them. Detection of chromosomal abnormalities at the embryonic stage during natural pregnancy is not possible. During IVF, the first stages of an embryo's life occur in vitro (outside the mother's body), so there is an opportunity to study chromosomes. This makes it possible to select embryos with a normal chromosome set for transfer into the uterine cavity, which in turn significantly increases the chances of a healthy pregnancy. It also significantly reduces the risk of genetic termination of pregnancy required.

It should be borne in mind that PGD does not completely replace prenatal diagnosis. It is known that chromosomal abnormalities are most often of maternal origin, in this regard, when performing PGD, it is effective to analyze not only blastomeres (embryonic cells), but also polar bodies (cells formed during the maturation of the egg)... It should be borne in mind that a woman's age over 45 is a risk factor for obtaining a small number of eggs and, accordingly, a small number of embryos. In this case, PGD may be ineffective and such factors serve to limit the use of PGD.

Male factor of infertility, namely severe spermatogenesis disorders such as oligozoospermia, teratozoospermia is also considered as an indication for PGD. It is believed that in this case, the number of spermatozoa with chromosomal abnormalities is increased, respectively, the risk of aneuploidy of embryos is increased.

The presence of chromosomal rearrangements (structural abnormalities) in the karyotype of one of the spouses, for example, translocations, is an indication for PGD... It is believed that such patients may have an increased risk of not only specific chromosomal abnormalities, but also the risk of aneuploidy for chromosomes not involved in rearrangement; therefore, in such cases, it is recommended to combine PGD on translocation with PGD on aneuploidy.

Patients can carry out preimplantation diagnostics in the IVF cycle at their own request without specific medical indications. Aneuploidy (abnormalities in the number of chromosomes) occurs in human embryos with a high frequency, even in normal conditions. A married couple can use the chance to reduce the risk of chromosomal abnormalities in embryos during IVF to reduce the risk of spontaneous abortion and the birth of a child with a chromosomal abnormality.

PGD ​​in translocations

Carriage of chromosomal abnormalities in the karyotype (for example, translocations, marker chromosomes) is a risk factor for chromosomal abnormalities in offspring. The risk is associated with the fact that unbalanced karyotypes are formed (with a lack or excess of genetic material), which can lead to spontaneous abortion or the birth of a child with a serious illness. Carriers of translocations are more likely to suffer from miscarriage in a married couple or infertility (especially if the carrier is a man).

In the presence of structural abnormalities in the karyotype of one of the spouses, it is recommended to carry out PGD to identify embryos carrying unbalanced chromosomal material. The selection of embryos with a balanced karyotype increases the chance of such a couple to become pregnant and give birth to a healthy baby by about 20 times.

PGD ​​for structural chromosomal abnormalities, in most cases with translocations, requires a preparatory stage, which is carried out before entering the treatment cycle. At this stage, a chromosomal disorder testing scheme is selected, which is individual for a specific translocation, and is tested on the blood cells of the parent carrying the genetic disorder.

FISH - a method for the study of unbalanced chromosomal abnormalities in embryos

FISH - (fluorescent in situ hybridization - fluorescent hybridization) - a method of genetic research in the case PGD ​​in translocations and also in the case PGD ​​numerical chromosomal abnormalities... For the study, special FISH probes are used - labels for certain sections of chromosomes associated with a specific fluorescent dye. FISH analysis identifies embryos with imbalanced disorders.

PGD ​​of monogenic diseases.

PGD ​​of monogenic diseases is recommended for families with an established genetic disease. Gene (monogenic) diseases are rare. But if the family already has a child with a gene disease, then the risk of giving birth to another child with the same disease in this married couple is high. The cause of a gene disease is a violation or mutation of a particular gene (part of DNA). Parents, being carriers of the mutant gene, are highly likely to pass it on to their offspring, which leads to the birth of sick children. Frequent gene diseases include cystic fibrosis, phenylketonuria, adrenogenital syndrome, Wernig-Hoffmann syndrome, hemophilia, sickle cell anemia. Preimplantation diagnostics allows selection of embryos without mutations and can be performed for gene diseases with different types of inheritance: autosomal recessive, autosomal dominant, sex-linked.

The indication for PGD is not any monogenic defect, but only a high genetic risk for a severe hereditary disease. In order for a married couple to plan PGD, both spouses must be examined for a gene defect - to have the results of a molecular genetic study in their hands, mutations must be identified. With an unclear nature of inheritance, an undetermined mutation, PGD planning is not possible.

Before carrying out PGD of a monogenic disease, it is necessary to carry out a preparatory stage during which a scheme for testing a genetic disorder is developed, which is individual for each pair.

Method for the study of gene defects in embryos

The main method of genetic research in carrying out PGD of monogenic defects is polymerase chain reaction (PCR). The PCR technique used in PGD is different from the conventional PCR techniques used in genetic analysis. The peculiarity consists in a small amount of material and a limited research time. During the preparatory stage, informative markers are identified and a scheme for the analysis of these markers together with mutations on single cells is developed. For genetic studies of the preparatory stage, the blood of both spouses is required, and the blood of relatives may also be required... What kind of biological material is required is determined by a geneticist. PGD ​​of a monogenic disease can be combined with the diagnosis of chromosomal abnormalities in embryos. With PGD of a monogenic defect, an analysis of the violation of a specific gene is carried out. But all human embryos are characterized by a rather high nonspecific risk of other genetic disorders - numerical chromosomal abnormalities (aneuplodia).

How is PGD done?

Preimplantation diagnosis is possible only within the IVF / ICSI treatment cycle... Preparation for the treatment cycle and the treatment cycle of IVF with PGD itself practically do not differ from the usual treatment cycle: a woman receives hormonal drugs to stimulate superovulation, transvaginal follicle puncture is performed, oocytes are fertilized with spermatozoa in an embryological laboratory. The embryo transfer is carried out on the 5-6th day. A special feature of the treatment cycle is that the ICSI procedure is used during IVF cycles with PGD. This is necessary due to the fact that a large number of sperm are added to the egg during conventional IVF. Then, when collecting polar bodies or blastomeres, there is a risk that the genetic material of a sperm that did not participate in fertilization will be included in the analysis along with the embryo cell.

Preimplantation genetic diagnosis is carried out after obtaining follicles and fertilization, during the development of the embryo in vitro, before transfer to the uterine cavity. The material for genetic research during PGD is the genetic material of the egg and embryo... The genetic material of the oocyte obtained in the IVF program is examined for polar bodies and blastomeres (an embryo of 6-8 cells) using an embryo biopsy.

It is believed that this procedure does not interfere with the further development of the embryo.

While genetic diagnosis is being performed, embryos continue to develop in an incubator in an appropriate nutrient medium until they are transferred to the uterine cavity on the 5th day of development. By this time, the embryo should have reached the blastocyst stage. Before transfer, the embryologist evaluates the structure and shape of the embryos (morphology). The result of genetic diagnosis is compared with the morphology of the embryos and a conclusion is made about which embryos are recommended for transfer to the uterus. The best morphological characteristics of embryos without genetic disorders are selected for transfer. Genetic research carried out within the framework of PGD is highly technological. The analysis is carried out on a very small amount of material - on biopsy polar bodies and blastomeres - and in a very short time. Only 2 days are allotted for the analysis of blastomeres. The embryo cannot continue its development outside the mother's body beyond the blastocyst stage (5th day after fertilization), therefore, the study must be performed within this short time.

For pre-implantation diagnostics, on the one hand, the latest, and on the other hand, proven methods of genetic research are used. For PGD of numerical chromosomal abnormalities, PGD in translocations, the FISH method (fluorescent hybridization) is used. When carrying out PGD of monogenic diseases - the PCR method.

Diagnosis of genetic disorders by polar bodies

When the female reproductive cell (oocyte) matures, it divides to distribute the genetic material. During this division, the oocyte distributes the chromosomal material 2 times. For the first time, chromosomal material (46 chromosomes) doubles and splits between two cells. One of these cells is called the first polar body. Then the second cell goes through another cycle of division, but without doubling the genetic material, and distributes its genetic material in equal amounts (23 chromosomes each) between the daughter cells. One cell becomes the egg, and the other becomes the second polar body. Polar bodies are characteristic of a mature egg and do not take further part either in fertilization or in the development of the embryo. These two polar bodies can be examined to determine whether the genetic defect is inherited by the egg, or whether the genetic defect has entered the polar body.

PGD ​​safety

Many future parents are concerned about the question: "Can PGD harm the child's health?" The blastomere is taken on the third day, when the cells of the unborn child are still universal, undifferentiated and some cells are replaced by others without harm to his health.... Polar body harvesting is considered even less invasive for the embryo. The risks of using PGD are mainly related to the general risks of the IVF program (for example, the risk of developing ovarian hyperstimulation syndrome).

Benefits of preimplantation diagnosis:

Selection and transfer to the uterus of only those embryos that do not have chromosomal abnormalities
Reducing the risk of having a baby with certain genetic defects
Reducing the risk of miscarriage (approximately 2 times)
Reducing the risk of multiple pregnancies (by about 2 times)
Increased chance of successful implantation (by about 10%)
Increased chances of a successful childbirth (by about 15-20%)

RISK during preimplantation diagnostics:

Risk of accidental damage to the embryo (<1%)
Erroneous diagnostics (up to 10%)
3.5% chance that an embryo with a pathology will be diagnosed as normal
10% chance that a healthy embryo will be diagnosed as an embryo with pathology
Cancellation of embryo transfer due to the fact that, according to the results of PGD, pathology will be detected in all embryos (up to 20%)

During pregnancy, it is possible to conduct diagnostic tests to determine whether the fetus is developing normally. Diagnosis of a number of pathologies is possible with the help of amniocentesis or chorionic villus sampling (sampling of fluid cells or tissues surrounding the embryo). Examination of the amniotic fluid and ultrasound examination can detect certain pathologies of the central nervous system of the fetus or other organs. You should discuss these tests with your obstetrician / gynecologist who monitors the pregnancy. These studies are not 100% reliable.

As in the case of natural conception, with IVF there is no guarantee that a child with undiagnosed physical or mental disabilities will not be born. The data available to date indicate that the risk of developing pathologies in children after in vitro fertilization (IVF) is not higher than that after natural conception.

For many parents, the birth of a child is a very desirable and long-awaited event. Especially if the couple cannot have offspring for a long time. In second place for mom and dad, after the fact of birth, is the good health of the baby. If during natural conception "ordering health" is problematic. Certain actions are required on the part of future parents, ranging from giving up bad habits, downloading examinations before pregnancy. But even this correct behavior does not provide the necessary guarantee. With IVF with PGD, it is easier to give birth to healthy offspring. The doctor checks their genetic information before. According to the results, only healthy embryos enter the uterus, and the bad ones are eliminated.

First, let's remember what is it? IVF is an in vitro fertilization procedure. It differs from the natural conception procedure in that the embryos are fertilized outside the woman's body and then placed inside the uterus. The success of IVF largely depends on the skill and experience of the doctors. Not on the last step in importance is the health of the expectant mother and genetic father.

PGD ​​at Eco what is it? Let's spell it out: preimplantation genetic diagnosis. With its help, the embryo is diagnosed even before it is transferred to the woman's body. PGD ​​is carried out in conjunction with the IVF program. Carrying out diagnostics, the doctor excludes in advance the presence of genetic pathologies in the unborn child. Since a practically healthy embryo is inserted into the uterine cavity, the risk of spontaneous miscarriage is reduced to a minimum value.

Under what conditions is it necessary to carry out PGD

Such a diagnosis is not always carried out. With it, the cost of IVF increases significantly. But under some conditions, it is mandatory. Let's list them:

1. The expectant mother is over 35 years old. The proportion of genetically normal embryos decreases with age, while the proportion of abnormal ones increases dramatically.
2. The genetic father is over 45 years old.
3. There has already been the death of children from an incompatible Rh factor.
4. Future parents have genetic diseases, that is, there is a serious risk of getting a genetically unhealthy baby.
5. The childless couple have already had several unsuccessful attempts at artificial insemination.
6. There were cases of spontaneous abortion for periods of 4-8 weeks.
7. The man showed serious spermatogenesis disorders or high abnormal sperm count.

The PGD procedure helps to identify a sufficient number of genetic diseases. By examining the chromosome set of the embryo, the doctor can detect more than 150 diseases. These include: Gaucher's disease, myodystrophy, achondroplasia, hemophilia, Gettington's disease, X-fragile chromosome, pemphigus, retinoblastoma, neurofibromatosis, Fanconi's anemia, Down's syndromes, torsion dystonia, polycystosis of the kidneys, myopathy, cystic fibrosis , phenylketonuria.

Diagnosis requires a significant number of follicles. Therefore, with PGD, a woman is given increased ovarian stimulation.

Objectives of PGD

If, according to the indications, the described diagnostics is necessary, then the doctors' actions will be aimed at achieving the following goals:

• A significant reduction in the risk of getting a genetically unhealthy child.
• in order to exclude the conception of a child with a disease associated with gender.
• Establishment of the Rh factor, which will help exclude hemolytic abnormalities in the embryo.
• Revealing information why implantation did not occur in previous IVF attempts.
• Immediately weed out embryos that are affected by incurable diseases.
• Selection of the necessary genetic information, which in the future will help the child to donate to a sick sister or brother.

How accurate is the PGD procedure?

Preimplantation diagnosis has proven to be accurate. The probability of a possible error does not exceed 5%. In addition, after the onset of pregnancy, a woman is prescribed and carried out combined screenings. If suddenly they give an unacceptable percentage, she undergoes invasive prenatal diagnostics.

The two sides of PGD: positive and negative

On the positive side, these are:

1. The ability to immediately establish and "order" the sex of the child.
2. Getting a healthy baby, thanks to replanting only good material.
3. It is very difficult to damage the embryo, the risk is less than 1%.
4. Possibility of preventing the conflict of razus factors.
5. Abortion is excluded due to identified genetic abnormalities.
6. The procedure is completely harmless to the embryo. At this stage of development, cells are completely interchangeable. The blastomeres taken for the procedure are easily replaced with new ones.
7. The success of the IVF program increases several times.
8. Congenital defects are almost completely excluded.
9. An excellent opportunity to help your own sister or brother, thanks to the selection of the necessary compatible material.

The advantages of the PGD procedure before IVF are impressive, but also significant. The negative side is manifested in the following factors:

1. Small, but still, the risk of damage to the embryo. You cannot get rid of it and it can come.
2. The likelihood of getting incorrect results. Sometimes healthy embryos are mistaken for unhealthy ones. They are weeded out to the side. It turns out that material with incorrect information is inserted into the uterus. To exclude such a negative factor, screening is carried out, which allows you to identify such information at a fairly early stage of pregnancy.
3. There is a 3-5% risk of getting an unhealthy baby.
4. The PGD procedure is very expensive.

PGD ​​price

Preimplantation genetic diagnosis costs a lot. The price range starts from 50,000 and ends at 130,000. The price includes expensive equipment and modern, expensive materials. The cost also depends on the number of embryos to be examined. Having decided on the PGD procedure, you can order an examination for one pathology or carry out diagnostics in full. If the procedure is performed by top-class specialists using professional equipment, the likelihood of the desired positive outcome increases.

The high cost of PGD can also be viewed from the other side. If there is a high risk of getting a genetically unhealthy baby, then it may be more likely to be born. An unhealthy child will need special care, expensive treatment, and general maintenance for life. Plus, the mental anguish of parents suffering for their child will always bother them. All of this in aggregate significantly exceeds the named cost of PGD. It turns out, after all, that it is better to carry out PGD.

PGD ​​methods

Doctors practice four methods of PGD. These are FISH, CGH, PCR, NGS. Let's consider them separately.

FISH method

It stands for fluorescent in situ hybridization. Or, in another way, it can be called the molecular-cytogenetic method. It is often used in the CIS countries. FISH is used for interphase and metaphase nuclei. The method allows you to identify the DNA sequence on the investigated chromosome. For the price, it is the most inexpensive diagnostic method. In terms of time, the study takes about 4-5 hours. FISH has a drawback: its method does not allow you to check all chromosomes.

The fish method also helps:

1. To carry out PGD of embryos that are carriers of chromosomal rearrangements.
2. Do a screening study for common chromosome aneuploidies.
3. Immediately determine the gender of the planned child.

CGH method (comparative genomic hybridization)

It can be used to check all chromosomes. The research takes quite a long period of time. It is expensive for the price. Its peculiarity: the resulting blastocysts are frozen, and further transfer is carried out using. In addition, this method checks embryos that have a strong potential for further implantation. Also, with CGH, the risk of misdiagnosis is significantly reduced, thanks to the biopsy of a large number of stem cells.

PCR method

It allows you to correctly diagnose diseases that arise due to dominant and recessive mutations (for example, diseases such as cystic fibrosis, hemophilia). PCR detects chromosomal abnormalities, determines antigens and Rh embryos. The PCR technique also checks close relatives. That is, in addition to diagnosing parents, they do a study of relatives for the presence of monogenic diseases. Such actions are necessary to detect gene abnormalities in embryos.

NGS method

Let's decipher: Now-Generation Sequencing. Translated as high throughput sequencing. This is the most innovative diagnostic. With NGS, the DNA sequence is checked, thereby accurately establishing the genetic pathology. The new technology is gradually replacing the first FISH technique and is widely used in foreign IVF centers. Practice has shown that success occurs in 70% of cases.

Preparing for PGD

If there is a need to combine IVF with PGD, then a geneticist will consult with the couple. Further, experts prescribe a full examination for future parents to identify all rearrangements and detect mutations. Further, accurate readings for PGD are established. At the last stage of preparation, the doctor explains the whole essence of the PGD program and describes the stages of the procedure.

Stages of PGD

Consider the stages of PGD:

1. The first stage begins with the standard IVF procedure. The process ends with a stage.
2. Embryological procedure of oocytes and sperm, micromanipulation. On the third day of culture, a biopsy procedure is performed. For this, one cell is taken from the embryo using a special microsurgical instrument. Biopsy uses mechanical, chemical and laser techniques. Next, the blastomere is fixed. In addition, biopsy of the trophectoderm and biopsy of the polar body of the oocyte can be performed. In the first case, 3-5 cells are taken from the trophoectoderm (usually they are on the outer layer of the blastocyst). A biopsy of the trophectoderm is performed on the 5th or 6th day and is a reliable option for identifying the karyotype of abnormal embryo development. Oocyte body biopsy is the best diagnostic option for female infertility. The genetic information in the polar body and in the nucleus of the egg is the same.
3. Hybridization with DNA probes. Probes are understood to be a special label. Each single chromosome has its own label. It is illuminated with color under the influence of fluorescent. Therefore, their identification is easy to carry out.
4. Obtaining visualized information on a computer monitor screen. Diagnostics of blostmasters that have been recorded. Obtaining final results by the 5th day of cultivation. Here doctors count the number of chromosomes. If their number corresponds to the accepted value, then the material is recommended for further transfer. If, instead of three chromosomes, two are observed or instead of the required two, one is visualized on the screen, then such material should not be transferred, it is abnormal.
5. Transfer of genetic healthy embryos on the 5th day of cultivation. This time is considered the most optimal. The material for use is presented in sufficient quantity and the biopsy procedure for the embryo is not so traumatic. If you transfer the material to the third day, then there will be a high probability of stopping the development process. In addition, in a three-day-old embryo, the genetic cells are very different. As a result, you can get false studies.
6. Carrying out after the transfer procedure.
7. Diagnosing a long-awaited pregnancy using a test (usually two weeks should pass after replanting).

Possible consequences of IVF with PGD

In addition to screening a pregnant woman after IVF, an analysis of the amniotic fluid and an ultrasound of the fetal nervous system are done.

Depending on the PGD method, the diagnosis will take from several hours to several days. The entire IVF cycle is 4-6 weeks. This interval also includes PGD.

The PGD procedure is performed on an outpatient basis. Sedation is only required when collecting eggs.

With IVF with PGD, the same risks arise as with IVF without PGD. It:

• Multiple pregnancy.
• Damage to the embryo at the stage of cell withdrawal.
• Reactions to drugs that can be used during the IVF procedure. These are mainly headaches, hot flashes and mood changes for the worse.
• There is a syndrome of ovarian hyperexcitation.
• Bleeding after oocyte retrieval.
• Sensitive spasms after the withdrawal procedure.
• The PGD study does not give a hundred percent result.
• Obtaining conception outside the uterus.

Contraindications

PGD, despite its necessity, efficiency and painlessness, has several contraindications for conducting:

1. The resulting embryo on the 2nd day of its development has less than 6 blastomeres.
2. Fragmentation of the embryo is greater than 30% (normal value is 25%).
3. The embryo contains multinucleated blastomeres.

Genetic diagnostics as a perfect method

Of course, a woman's biggest fear is to give birth to a genetically unhealthy baby. The PGD method allows you to weed out genetically diseased embryos, thereby immediately giving the child health. Parents do not have to fear for the health of long-awaited children. If someone believes that genetic research makes a child according to a template, then this is not the case. You cannot create a design project for an unborn baby. The character and personality do not depend at all on the genes obtained. Even artificially cloned animals are not alike. Genes are responsible for external data.

IVF together with PGD strongly influenced the verdict “sterile”. More precisely, this is no longer a sentence, but a special disease that can be easily bypassed with the help of IVF. The method allows you to overcome a number of health difficulties, namely:

• Obstruction of the fallopian tubes.
• Lack of fallopian tubes.
• Depletion of the ovaries.
• Lack of a long time of the ovulation process.
• Disruption of the ovaries.
• Adhesions in the small pelvis.
• Lack of sex glands.

Even if a woman does not have a uterus, the problem can still be solved. But at the same time, her ovaries should function normally. To do this, eggs are taken from a woman, fertilized with the sperm of her husband, and the embryo is placed in the body of another woman (for example, a relative or a surrogate mother).

PGD ​​is an absolutely safe procedure. The diagnostic procedure is done as soon as possible, when a full-fledged organism can be obtained from each taken cell. There will be no harm to the health of the embryo, respectively, the unborn child will also be healthy.

Genetics continues to evolve. It allows you to get healthy offspring and future parents in advance, who are more likely to be free of genetic diseases. An adult realistically estimates the future, he wants to give the baby a full life in advance and in the future to get offspring (grandchildren) from him. Genetics certainly helps with this.

So, the method of preimplantation genetic diagnosis is an addition to the entire IVF program. Do not pay attention to the cost of the procedure. If money has already been found for the IVF itself, then the rest of the necessary funds are minuscule. Child health is not worth the savings.

PGD ​​- all pros and cons for a successful pregnancy: preimplantation genetic diagnosis of embryos

Should you do PGD to increase your chances of a successful pregnancy? The decision to conduct preimplantation genetic diagnostics causes a lot of controversy on the forums of future IVF moms. The relevance of the issue remains at its peak, especially since the cost of PGD in IVF is increasing. So does the analysis affect the likelihood of pregnancy?

• Objectives and Benefits of Analysis
• Why do PGD in IVF?
• Indications for conducting
• Diagnose or not?
• How is the research done?
• Stages
• Canceling the transfer after diagnostics
• Preparation for the program
• What should be done when pregnancy occurs?

Objectives and Benefits of PGD Analysis

Preimplantation genetic diagnosis of PGD with IVF is a method for examining the genome before transferring the embryo into the uterine cavity. The main goal is to select an embryo for replanting without genetic pathologies, which are carried by the parents.

For a successful pregnancy, it is important to transfer not only a high-quality, but also a genetically "healthy" embryo. The main advantage of PGD is a reduction in the risk of both spontaneous abortions and artificial termination in case of pathology detection at the perinatal stage (i.e., when pregnancy has already occurred).

Why do the analysis?

The positive effect of PGD is quite weighty. Why is it recommended to conduct an analysis? The analysis is important because it:

• increases the percentage of implantation;
• reduces the rate of spontaneous abortion;
• reduces the birth rate of children with genetic abnormalities (the most common pathology is the birth of a child with Down syndrome);
• makes it possible to choose one, but the best embryo for transfer to the uterus (avoids);
• promotes the birth of a healthy full-term baby on time;
• allows you to select the gender of the child, which is necessary to prevent gender-linked genetic abnormalities.

The disadvantage of PGD is that the cost of the ART program rises.

Indications for PGD in IVF

PGD ​​is shown:

• married couples with a high genetic risk (family cases of genetic diseases);
• women over 35. With age, the number of genetically abnormal embryos increases significantly, while the proportion of normal ones decreases.
• married couples whose partner's age is over 45;
• if a man has severe spermatogenesis disorders, a high percentage of abnormal sperm, or single germ cells.
• women with, especially in early pregnancy from 4 to 8 weeks.
• couples after multiple.

To conduct a PGD analysis or not?

PGD ​​is also relevant for couples who have undergone, but have not become pregnant. One of the possible reasons is a genetic disorder.

An embryo of normal quality may have an abnormal genotype. Good quality of the unit to be carried is not always a good quality genotype.

Therefore, PGD in IVF is always the second stage in the selection of embryos for transfer into the uterine cavity. The first stage is extended cultivation to the blastocyst stage. The first three days, the development of the embryo takes place on the mother's reserves. On the 4th day, its own genome is included in the work. Gross violations of genetic information in cells block further development.

How common are genetic abnormalities in embryos?

Embryos of excellent and good quality have a fairly large percentage of karyotype changes (aneuploidies), which do not allow normal development in the future. The data in numbers can be found in Table 1.

Table 1. The quality of blastocysts and the quality of the genetic set of chromosomes

It can be concluded that in 25–39% of cases, pregnancy can produce embryos of satisfactory and unsatisfactory quality, since they are genetically completely “healthy”. But in order to identify them, you need a PGD analysis. Diagnostics on such embryos is extremely difficult. There is too little cell material and it is difficult to obtain it without harming the developing organism. The risk of stopping the development of embryos of satisfactory and unsatisfactory quality after biopsy is very high. Most often they are not examined.

PGD ​​method - how is the study done?

There are 4 methods of conducting PGD in the world, which require different equipment of the genetic laboratory:

• Fluorescent in situ hybridization - FISH. The most accessible and most often used method in the CIS countries. Compared to others, it is inexpensive, requires little time - only 4–5 hours. Its disadvantage is that it does not allow examining all chromosomes.
• Comparative genomic hybridization (CGH). The method allows examining all chromosomes; it is quite expensive and time-consuming. When using it, blastocysts are frozen and transferred in a cryoprotocol.
• Polymerase chain reaction (PCR). Allows to investigate monogenic chromosomal mutations and requires preliminary diagnosis of the parents. First, a survey of the married couple is carried out, the presence of a mutation is revealed (determined in which gene). And only after it was detected in the parents, the mutation is looked for in the embryo.

• Now-Generation Sequencing is a PGD method that allows you to study all chromosomes and polymorphisms (complete genomic analysis), very expensive. Today it is one of the high-resolution technologies that allows you to study not only the chromosome set, but also mutations in any of the chromosome regions.

Stages of PGD

Investigate the genotype in three-day and five-day embryos. Recently, the trend is towards the fact that the analysis is more often performed on 5-days. The reason is the possibility of a more accurate analysis based on the results of the study of not one cell, but several. For PGD embryos on the third day of cultivation, 1 (maximum 2) blastomeres are taken. When analyzing blastocysts, 5-7 cells are taken from the trophectoderm (the area of ​​embryonic tissue from which the placenta develops). The cells from which the actual organs and tissues of the child develop are not involved in the study.

Stages of PGD:

1. Embryo biopsy.
2. Fixation of cell nuclei on a slide (slide for work under a microscope).
3. Hybridization with DNA probes. The probe is a label, each chromosome has its own label, which differs in the color of the fluorescent glow. Each pair of chromosomes glows with a specific color, which allows them to be identified.
4. Visualization on a computer monitor of a fluorescence microscope and interpretation of the information obtained. In each nucleus, the number of chromosomes is counted. If the number of chromosomes is normal, then such a blastocyst is recommended for transfer. If trisomy (three chromosomes instead of two) or monosomy (one chromosome instead of two) is detected, the embryo is not recommended for transfer. He's abnormal.

On which day is it better to carry out PGD on 3 or 5?

Recent studies have shown that embryo biopsy for PGD on the third day carries a high risk of stunting. Plus, on the 3rd day, the embryo has a high level of mosaicism (the cells are genetically different), which leads to the appearance of false results and increases the percentage of errors.

PGD ​​of the embryo on day 5 is widely used, since a biopsy is less traumatic for it, and there is more material for research.

Other benefits of PGD for successful IVF:

• blastocysts are easier to tolerate a biopsy;
• higher pregnancy rate;
• low risk of developmental arrest.

There is a concept of "complete blastocyst" - this is when complete differentiation of embryonic cells has occurred and the intracellular mass and trophectoderm are clearly visible under the microscope. It is these blastocysts that undergo preimplantation genetic diagnostics.

There is a concept of "early blastocyst". Their differentiation has not yet been completed. Such embryos are not biopsied, but continue cultivation up to 6 days, and after the completion of differentiation, PGD is done.

Cancellation of transfer after PGD

The percentage of cancellation of embryo replanting after PGD is quite high, and its level increases with the age of the woman. It often happens that after the examination there is no embryo suitable for transfer. In women over 40, the embryo cull rate is 25%.

One out of four women, after PGD examination, have no genetically quality embryos at all, and no transfer is carried out.

PGD ​​risks

The examination is invasive, therefore there are risks for the embryo itself and risks of inaccuracy of the survey data obtained:

• the risk of damage to the embryo during biopsy, especially for 3 day old embryos;
• the risk of biological error with a high level of mosaicism;
• risk of equipment malfunction or insufficient doctor's qualifications.

Preparing for the IVF program with PGD

Before the IVF program with PGD, a consultation with a geneticist is required.

Preparation for IVF + PGD includes:

• examination of the couple (karyotyping, identification of mutations, rearrangements);
• determination of indications for PGD;
• clarification of the essence of the program, possible risks and errors, the accuracy of the method.

It is also important to discuss with the doctor the tactics of behavior in the case of a small number of embryos (it is not advisable to carry out PGD on one embryo). And also actions in the absence of embryos for transfer according to the data of genetic research.

When pregnancy occurs after successful IVF with PGD

With a successful pregnancy after IVF with PGD, a woman is expected to be closely monitored with ultrasound screening, perinatal screening in the first and second trimester, as in a natural pregnancy.

When pathology or abnormalities are detected, a decision is made on the need for invasive perinatal diagnostics. This is a rather complicated procedure, it is equivalent to an operation and is performed under sterile conditions. Its purpose is to obtain a biopsy material - chorionic villi, placenta or amniotic fluid for analysis. The samples obtained are examined and diagnosed.

In the case of spontaneous abortion, a genetic examination of the abortion material is carried out.

Standard diagnostic kits

Standard kits are widely used for research. With their help, you can diagnose violations in 3 chromosomes (X, Y and 21 pairs), in 9 (13, 18, 21, 15, 16, 17, 22, X, Y) or you can examine 24 chromosomes. If necessary, the clinic orders an individual set for PGD diagnostics from the company to the manufacturer.