Fertility diagnosis

From the initial infertility investigation, diagnosis and treatment, our goal is one, to help you achieve your goal of completing your family.

Infertility can be caused by both the man and the woman, or a combination of both. That is why it is very important that the approach to infertility assessment always incorporates both members of the couple.

At the initial appointment, the goal is to accurately define the possible cause of infertility and create an effective treatment plan. The first step is a comprehensive history which is supplemented by diagnostic tests.

The following is a description of the most common diagnostic tests in relation to female and male infertility:

Female Fertility Investigation

The assessment of ovarian function and egg reserves is a very important element in the initial assessment of infertility. The detection of reduced oocyte reserves during the initial investigation of infertility determines the choice and type of infertility treatment, which is individualised according to the specificities and needs of each couple and includes :

  • Checking the FSH, LH and estradiol hormones on the 2nd or 3rd day of the period.
  • Anti-Mullerian Hormone (AMH). This test can be done on any day of the cycle.
  • Ultrasound scan of the ovaries to measure follicle diameter 2 -10 mm (AFC) and ovarian volume.

If there is evidence of premature ovarian failure further investigation and genetic testing is recommended.

Testing of hormones that affect reproductive function such as thyroid or prolactin tests are also included in the initial investigation.

  • An ultrasound scan of the uterus gives us information about the size and texture of the uterus.
  • Hysterosalpingography is the initial test that is usually done as a first step in assessing the patency of the fallopian tubes. This test also gives a more detailed view of the uterine cavity and is done in the first half of the cycle, usually from day 8 to day 12.
  • Saline-Infusion Sonography (SIS) is a new ultrasound technique that allows us to check and accurately diagnose if there are polyps or fibroids protruding into the intrauterine cavity.
  • Hysteroscopy or Laparoscopy. If indicated, it provides the advantage of being able to directly view the anatomy of the intrauterine cavity, or fallopian tubes as well as the other pelvic organs.

Assisted reproductive treatments

The monitoring of ovulation in a woman’s cycle is carried out by ultrasound scans and measurements of the hormones oestradiol (E2) and LH in the blood, so that the day of ovulation can be determined very accurately.

The couple therefore have planned intercourse on the exact day the woman ovulates.

It is a simple assisted reproduction treatment, which is mainly applied to couples with unexplained infertility. It is divided into homologous when using the sperm of the spouse or partner and heterologous when using third donor sperm.

  • Homologus Insemination

    Necessary conditions for the treatment of insemination to have high pregnancy rates are that the woman must have healthy and heavy fallopian tubes and the sperm of the husband/partner must have good enough parameters.

    Insemination can be performed in the woman’s natural cycle, however in order to increase the success rates, we induce mild drug stimulation of the ovaries, with the aim of producing 2 to 3 follicles.

    The cycle is monitored by ultrasound scans and blood measurements of the hormones estradiol and LH. This is how we check that the follicles are developing properly.

    On the day of ovulation, which is either precisely detected by checking endogenous hormones or induced by medication, the sperm is specially processed and then placed in the uterus with a thin, flexible catheter. This increases the chances of the sperm reaching the fallopian tubes, where fertilisation of the egg normally takes place.

    Statistically, treatment with intrauterine insemination has better results when the woman is under 38 years old.

    Heterologus Insemination

  • If the spouse/partner cannot produce any sperm, then donor sperm is used.

    Treatment of heterologous insemination in women is done in exactly the same way as in homologous insemination.

    On the day of ovulation, 1 to 2 ampoules of frozen donor sperm are thawed from the sperm bank and placed directly into the uterus.

    See also:

    Sperm Donation & Legal Framework

This treatment is mainly used in women in whom the use of ovarian stimulating drugs is contraindicated, such as when the ovaries do not respond to these drugs and therefore cannot produce eggs.

Another reason is when it is contraindicated to increase the hormone estradiol, such as if there is a history of breast cancer. Estradiol is the hormone produced by the follicles. The administration of stimulation drugs (FSH) causes multiple follicles to grow, resulting in a significant increase in the levels of estradiol in the body. When we want to avoid this, we perform IVF treatment in the woman’s natural cycle.

We monitor the growth of the follicle that the woman produces during her natural cycle with ultrasounds and blood tests.

In the ovulation phase, we take the one egg that is produced naturally and fertilize it with the husband’s sperm.

The resulting embryo is placed in the uterus, just as embryo transfer is done in the pharmaceutical cycle.

In IVF treatment with the help of medication, the ovaries are stimulated to mature and receive more than one egg. The reason for administering these drugs is to induce the development of more follicles, in which the eggs grow and mature. The main aim of the medication, is to increase the success rates, as when there are more eggs, there is a greater chance that there will be at least one good quality embryo to transfer into the woman’s womb to achieve pregnancy

Pharmaceutical Ovarian Stimulation 

The drugs used are FSH, drugs that are very similar to the hormone that is naturally produced by the pituitary gland and is necessary for the development of follicles and eggs in women.
Because the treatment is individualised, the regimen – protocol for taking medication – is chosen and used accordingly. While taking them, the woman is monitored by ultrasound and measurement of the hormone estradiol in the blood, so that everything is under control and at the right time we can take the eggs.
The ovulation procedure is performed by administering a light sedation intravenously, so that it is completely painless.
The oocytes are collected transvaginally by ultrasound examination. It takes between 10 and 20 minutes, depending on the number of follicles.
On the day of the egg collection, the husband/partner donates the sperm, which will then fertilise the eggs.
After ovulation, the woman sits in the clinic for about 1 hour and then can return home.
What is Fertilization
Fertilisation is a series of actions that start with the entry of the sperm into the egg. The activation of the ovum, the completion of Metaphase II, the exit of the second polar particle and the fusion of the ovum and the spermatozoon lead to the formation of pronuclei containing the genetic material from the father and the mother, 23 chromosomes each. Fertilisation is completed by the fusion of the 2 pronuclei, thus creating the normal number of chromosomes in the embryo, i.e. 46 chromosomes.
Fertilisation in the Laboratory
Immediately after receiving the eggs, they are evaluated and prepared for fertilization. Only the initial stage of the fertilisation process takes place in the laboratory. The fertilization process is completed by the egg and sperm in special ovens under completely controlled conditions.
Fertilization in the laboratory can occur in two ways. Depending on the parameters of the sperm and the couple’s medical history, either classical IVF or ICSI is chosen.
  • Classic IVF

    In simple fertilisation, the eggs are placed together with the sperm, which has been properly prepared, in the dishes where they remain for 16 to 20 hours in controlled laboratory conditions, the so-called ovens, in order to achieve fertilisation.

  • Microfertilization – ICSI (intracytoplasmic sperm injection)

    Microfertilisation, often referred to by its initials ICSI, was developed in 1992. It is a technique performed in a controlled laboratory environment and is now part of IVF treatment. A morphologically normal sperm cell is inserted directly into the cytoplasm of a mature oocyte, thereby penetrating the transparent zone and the cell membrane of the oocyte.

    Indications for the use of microfertilisation

    • Low sperm count per ml (oligospermia)
    • Low sperm motility (asthenospermia)
    • High percentage of spermatozoa with abnormal morphology (teratospermia)
    • Sperm obtained by a surgical technique
    • Problems with fusion or problems with penetration of the sperm into the egg
    • History of fertilisation failure or low fertilisation in a previous IVF attempt
    • Unexplained infertility
    • Frozen semen in small quantity
    • When Preimplantation Genetic Diagnosis of Embryo is to follow
IMSI Intracytoplasmic Injection of Morphologically Selected Sperm Into the Ovum
The use of microscopes with special lenses allows a higher magnification (x 6000) of the spermatozoa. It is a technique that allows to select more precisely the most normal spermatozoa to be used to fertilise the eggs.
According to the international literature, selection of spermatozoa by the IMSI method shows better rates of achieving pregnancy.
Embryo transfer
Embryo transfer is usually done 3 to 5 days after egg collection and is a simple and painless procedure.
The embryos are placed with a thin and flexible catheter through the cervix into the uterus.
It is carried out by ultrasound scanning, which allows us to identify the best place for their placement.
14 days after ovulation, a pregnancy test (detection of chorionic gonadotropin in the blood)

A blastocyst is an embryo that has continued its cell divisions and has reached the 5th day of development. At this stage, it has 50 to 200 cells, swells and increases in size, forms a cavity inside and is ready to be hatched from the transparent zone and implanted.

The advantages of culturing embryos to the blastocyst stage are:

on day 5 there is better synchronisation between the endometrium and the embryos
the endometrium under the influence of progesterone shows less contraction
the embryos that have continued their development up to the blastocyst stage have shown to have better growth potential and this allows us to select the best embryos for embryo transfer.

The disadvantages are:

Probably not all embryos will continue to develop to the blastocyst stage when outside the woman’s body, so some embryos will develop better if transferred to the uterus in a shorter time frame.
Freezing embryos at this stage is done on a smaller number of embryos.

In cases in which the man has a complete absence of spermatozoa during ejaculation (azoospermia), there is the possibility of obtaining sperm from the epididymis or from the testicles by surgery by a urologist.

They can be obtained in two ways directly, either by needle aspiration (FNA) or by biopsy in which small pieces of testicular tissue are taken.

Azoospermia, i.e. the absence of sperm from the semen, is divided into obstructive and non-obstructive azoospermia.

Obstructive azoospermia

There is a lack of sperm from the semen, but the production of sperm by the testicles is normal. In this case the spermatozoa do not appear in the semen due to obstruction of the seminal ducts or even in congenital lack of them.

Non-obstructive azoospermia

In the case of obstructive azoospermia, the lack of spermatozoa is due to an inability of the testicles to produce them. In 50% of men who have non-obstructive azoospermia, there may be some areas in the testicle that produce sperm, but they do not appear in the semen obtained at ejaculation, although there is no obstruction of the seminal ducts. In these cases, a biopsy taken directly from the testicles enables us to find spermatozoa and use them to fertilise the eggs.

The sperm obtained in this way have very low mobility and that is why microfertilisation (ICSI) is always performed.

It is possible to freeze the testicular tissue and store it for use in subsequent treatment cycles.

The freezing of sperm and testicular tissue enables their storage and use in possible future IVF treatment.


  • When the number of sperm in the semen is very low and there is a chance that it will be zero in the future, the man has the ability to store his genetic material.

  • In some cases, men of childbearing age may need to undergo treatments that may damage spermatogenesis, such as chemotherapy. In these cases, their sperm may be frozen before undergoing these treatments.

Since 1983, when the first baby was born from frozen embryos, embryo cryopreservation has been an important part of assisted reproduction.

The ability to freeze and store embryos increases the potential for conception with IVF treatment, while reducing the unused and lost embryos created by this treatment.

Embryos that have been frozen and stored can be used in another cycle of treatment.

However, of the embryos subjected to the freezing process – at -196°C – there is a possibility that not all of them will survive. Statistically, about 80% of embryos survive this process of freezing and thawing.

In recent years, various reasons, mainly social, have led women to delay having children. However, fertility is directly linked to a woman’s age. And while fertility is highest between the ages of 20 and 28, by the age of 35, the chances of conception per month are halved, and by the age of 40, they fall even further. Oocyte cryopreservation offers women a very important opportunity, that of preserving the eggs at the age of the woman at which they were cryopreserved. That is, while the chronological age of the woman when that woman wants to use them and have a child will have increased, the biological age of the eggs will have remained the same as when they were cryopreserved.

In addition, this technique can also help women in preserving their fertility when these women are about to undergo any of the treatments that usually cause destruction of the follicles and eggs in their ovaries, such as radiotherapy or the use of chemotherapy drugs.

A woman conceives and gives birth, following IVF and transfer of fertilised eggs (embryos), to a woman other than herself on behalf of another woman who wishes to have a child but is unable to carry it herself for medical reasons. The woman who conceives and gives birth is called the ‘gestational mother’ and the second woman is called the ‘birth mother’. Surrogacy is permitted with a court-ordered licence granted before the transfer of the embryos. Court authorisation is granted on application by the woman who wishes to have a child, if it is proven that she is medically unable to bear a child.

See also:

Legal framework – Surrogacy

Specialised genetic tests


PGD can be realised in one of the 3 stages of development:

Biopsy of embryo blastomeres
A PGD embryo blastomere biopsy is performed by removing 1 or 2 cells (blastomeres) from the embryo at the 6-8 cell stage on the 3rd day of development.

Biopsy of cells from the trophoblast when the embryo is at the blastocyst stage.

Biopsy of the polar particle of the ovum
By removing the first or second polar particle, or both, the chromosomes of the ovum are diagnosed before fertilisation.

While polar body biopsy is an indirect way of genetically testing the chromosomes of the ovum, embryo biopsy allows the detection of genetic abnormalities from both the mother and the father.

The genetic material of the cells taken from the oocytes or embryos is tested for the presence of genetic abnormalities.

Depending on the type of diagnosis required, the corresponding technique is applied.

Comprehesive Chromosome Screening

Nowadays, new techniques allow us to analyse the whole chromosome, and give very high implantation rates of healthy embryos. With this technique, the healthiest embryos in the group can be selected for transfer.

This technique is not suitable for all couples. However, they may benefit from the Comprehensive Chromosome Screening technique:

  • Women over 35 years of age.
    It is known that embryos from older women are more likely to have chromosomal abnormalities. It is believed that most of the time embryonic factors are responsible for embryos not implanting and not giving birth. And as the age of the woman increases, so does the number of embryos that have chromosomal abnormalities. Studies conducted on women aged 35 to 39 show that 20% of embryos have chromosomal abnormalities. This percentage increases to over 50% when women are 40 years old and over. These embryos will either not implant at all because of their chromosomal abnormalities or will end up in miscarriage.
  • Women with a history of miscarriage.
  • When infertility is due to a severe male factor where there are very low sperm parameters.
  • Where there are multiple previous failed implantation attempts in IVF treatment.

PCR – polymerase chain reaction 

It is a technique that allows the amplification of copies of specific regions of DNA and is used to detect mutations in a particular gene.

FISH – Flurorescence in-situ hybridization 

It is used to diagnose numerical or structural chromosomal abnormalities and to determine the sex of the foetus for the diagnosis of sex-linked diseases.

It is a test where the chromosomes in the embryos are analysed before they are transferred to the womb. PGS was developed to improve embryo selection in couples undergoing IVF treatment due to infertility, believing that if the genetic status of the embryos can be determined, it will lead to a higher implantation rate per embryo and increase pregnancy rates.

To date, embryos selected for transfer during IVF treatment are chosen based on their morphology. Unfortunately, however, the morphology of the embryos does not confirm how healthy these embryos are. Pre-implantation genetic testing makes it possible to transfer embryos that have an even number of chromosomes. These embryos have a higher implantation rate, lower miscarriage rate and the pregnancy has a higher chance of making it to term and having a healthy baby.

Πρόκειται για μια εξέταση κατά την οποία αναλύονται τα χρωμοσώματα στα έμβρυα πριν αυτά μεταφερθούν στη μήτρα. Η PGS αναπτύχθηκε για να βελτιώσει την επιλογή εμβρύων σε ζευγάρια που υποβάλλονται σε θεραπεία εξωσωματικής γονιμοποίησης λόγω υπογονιμότητας, πιστεύοντας ότι αν μπορεί να προσδιοριστεί η γενετική κατάσταση των εμβρύων, αυτό θα οδηγήσει σε υψηλότερο ποσοστό εμφύτευσης ανά έμβρυο και θα αυξήσει τα ποσοστά εγκυμοσύνης.

Μέχρι σήμερα, τα έμβρυα που επιλέγονται για μεταφορά κατά τη διάρκεια της θεραπείας εξωσωματικής γονιμοποίησης επιλέγονται με βάση τη μορφολογία τους. Δυστυχώς, όμως, η μορφολογία των εμβρύων δεν επιβεβαιώνει πόσο υγιή είναι αυτά τα έμβρυα. Ο προεμφυτευτικός γενετικός έλεγχος καθιστά δυνατή τη μεταφορά εμβρύων που έχουν άρτιο αριθμό χρωμοσωμάτων. Αυτά τα έμβρυα έχουν υψηλότερο ποσοστό εμφύτευσης, χαμηλότερο ποσοστό αποβολών και η εγκυμοσύνη έχει μεγαλύτερες πιθανότητες να φθάσει μέχρι το τέλος της και να αποκτήσει ένα υγιές μωρό.

This is a test in which the chromosomes in the embryos are analysed before they are transferred to the womb. PGS was developed to improve embryo selection in couples undergoing IVF treatment for infertility, believing that if the genetic status of embryos can be determined, this will lead to a higher implantation rate per embryo and increase pregnancy rates.

To date, embryos selected for transfer during IVF treatment are chosen based on their morphology. Unfortunately, however, the morphology of the embryos does not confirm how healthy these embryos are. Pre-implantation genetic testing makes it possible to transfer embryos that have an even number of chromosomes. These embryos have a higher implantation rate, a lower miscarriage rate and the pregnancy has a better chance of going to term and having a healthy baby.

  • Peripheral blood karyotype

  • Cystic fibrosis mutation analysis

  • Detection of microdeletions on the Y chromosome

    Karyotyping is a test that allows us to identify and assess the number and structure of chromosomes. Chromosomes contain the genetic information from DNA and are located in the nucleus of every cell in the body.

    Normally there are 23 identical pairs of chromosomes in each cell, a total of 46 chromosomes. Only eggs and sperm have 23 chromosomes each.
    During fertilisation, when the egg and sperm fuse, the embryo is formed, which consists of 46 chromosomes, i.e. 23 from the mother and 23 from the father.

    When the mother or father in a couple has an abnormal karyotype, this abnormality will be passed on to the eggs or sperm respectively and is therefore inherited by the embryos created. The result of this is constant implantation failures or even constant miscarriages.

    The chances of a couple with infertility problems having an abnormal karyotype are around 4%.

    In the context of assisted reproduction, the karyotype test is carried out by taking blood samples.

Male Infertility Investigation

The spermogram is done according to WHO (World Health Organization) criteria, is an essential test in the initial investigation of a couple’s infertility and gives information on both the quantity and quality of sperm and spermatozoa.

However, semen parameters are not always related to the fertility of the sperm and therefore it is impossible to predict the chances of pregnancy based on the results of the semen analysis.

Depending on the case, genetic tests such as the following may be indicated

Apoptosis , the programmed cell death of spermatozoa, is characterised by fragmentation of sperm DNA. High rates of DNA fragmentation affect the development of embryos and are associated with reduced chances of success because these embryos cannot develop and give birth to a healthy baby.

Samples in which only up to 15% of the sperm have fragmented DNA are considered to have a high potential to form viable embryos, but the chances decrease when the percentages of fragmented sperm DNA increase. High percentages of fragmented DNA – >29% – affect embryo quality and development.

Molecular cytogenetic testing examines the percentage of aneuploidies (numerical chromosomal abnormalities caused during cell division of meiosis) in spermatozoa.

The percentage of aneuploidies in the sperm, normally, should not exceed 5%.