Research into infertility

In this part of the site we outline the wider context of assisted fertilisation:

  • how does a person become pregnant in a natural reproductive cycle?
  • what exactly is reduced fertility?
  • what research can we do at the CRG to find possible causes?

Tests for men

During the first consultation at the CRG the doctor will find out what tests are necessary based on an interview.

What you can do to make sure this consultation is as efficient as possible?
More information is available here?
  • In the overview: a list of all examinations the CRG performs or can have performed as part of your fertility treatment;
  • in "looking for the cause" more information about the why of the tests; and
  • in the pages after that, more information about every specific examination.

Current tests
Anamnesis (medical history)
at consultation
Physical examination
at consultation
Routine sperm analysis
sperm sample | in lab
Hormonal profiling blood test | in lab
Tests based on specific indications
Biochemical sperm components (fructose and alpha-glucosidase)
sperm sample | in lab
Hypo-osmotic swelling test (HOS-test) sperm sample | in lab
Mixed anti-globulin reaction test (MAR-test) sperm sample | in lab
Tray-agglutination-test (TAT) blood test | in lab
Cytogenetic testing blood test | in lab
Microdeletion testing blood test | in lab
CFTR-gene mutation test (cystic fibrosis gene) blood test | in lab
Transrectal ultrasound (TRUS) - urology department image | outpatient
Scrotal ultrasound - urology department image | outpatient
Testicular biopsy day admission
Testicular sperm aspiration outpatient or day admission
Additional tests
Sperm survival test
sperm sample | in lab
FISH-analysis of sperm sperm sample | in lab
Electron microscopy sperm sample | in lab

Blood analysis

Every fertility treatment is preceded by a series of blood tests for men (see practical aspects):
  • to establish detailed information regarding hormonal balance;
  • to determine the structure and number of chromosomes; and
  • to detect antibodies (infection test) against jaundice (hepatitis B and C), syphilis, the HIV virus (AIDS) and sperm.

There are many causes of male infertility.

Pre-testicular causes

This implies imbalances in the hormonal impulses which come from the brain or adrenal glands, which also play a hormonal role.
An example of hormonal imbalance is the 'Kallman syndrome': an absence of certain structures in the hypothalamus, making the production of GnRH impossible. Men with this condition have no sense of smell either.

Testicular causes
Refers to problems located in the testicles, where sperm is made.
The problem can originate - although not genetically determined - from the development of the testicles, or at a later stage. We refer to the latter as 'acquired abnormalities' (e.g. varicose veins).
    Post-testicular causes, ‘past’ the testicles
    These problems can be:
    • physical', e.g. obstructions in the epididymis or vas deferens) and not genetically determined:
      • a non-genetic example is a blockage of the epididymis after an infection;
      • a genetically determined abnormality may include an absence of the vas deferens at birth.
    • but they can also be 'functional', such as impotence.

    Other causes
    E.g. infectious or immunological. In the latter case the man produces antibodies against his own sperm. For the former see e.g. STDs and infertility.
    (In)fertile?
    Normozoospermia
    Azoospermia
    From 5 million (and less) sperm per ml. the man has strongly reduced fertility. See the sperm quality.
    An ejaculate of at least 2 ml, containing a minimum of 20 million sperm cells per ml, or a minimum total of 40 million sperm cells, of which 50% are motile and at least 30% are of normal shape.
    Absence of sperm in the ejaculate.
    check that the penis is normal;
    whether both testicles have descended and are of normal size and there are no growths;
    the structure of your sex organs will be examined;
    the vas deferens will be palpated to check for abnormalities;
    the epididymis will also be checked that it is (fully) intact and if so,
    whether it is not hardened or shows small cysts.
    During your first consultation appointment at the CRG you will talk to a doctor who is specialised in fertility medicine. The consultation consists of a full medical history and possibly a physical examination.

    Preparation for the consultation  

    To efficiently facilitate your first consultation at the CRG,
    • you will be asked to complete a detailed questionnaire beforehand. You can download it here;
    • if you have ever had previous related treatment (e.g. at the urologist), bring your medical file; and
    • any results of already performed tests.
    If you prefer not to do this yourself you can also ask the CRG doctor to request your medical file.

    Medical history

    The doctor attempts to gather as much relevant information as possible regarding your medical history, current physical condition and health status.
    Many of the questions the doctor will ask you will be of an intimate nature. They are all aimed at identifying the reasons why you and your partner are having difficulties in conceiving.
    Below are a few examples of questions you can expect:
    • whether either of you have any children from a previous relationship;
    • how long you and your partner have been trying for a baby;
    • the woman's age;
    • whether you have regular penetrative sex with ejaculation;
    • how frequently you have intercourse; at which stage of the womans' cycle;
    • whether you experience any problems whilst having sex;
    • which sorts of contraception you have ever used and your experiences with them;
    • whether you have ever had surgery;
    • dietary habits, alcohol use and other lifestyle issues;
    • your family's medical history. This is to detect any possible hereditary conditions. During your first consultation appointment at the CRG, your doctor will take a full medical history to have a complete medical picture of your general physical status and state of health. This is followed by a physical examination, assuming that one has not been performed recently by another doctor.

    Physical examination      

    The history can be followed by a physical examination, unless another doctor recently performed one.
    For a man the physical examination will include the following:

    • check that the penis is normal;
    • whether both testicles have descended and are of normal size and there are no growths;
    • the structure of your sex organs will be examined;
    • the vas deferens will be palpated to check for abnormalities;
    • the epididymis will also be checked that it is (fully) intact and if so,
    • whether it is not hardened or shows small cysts.

    If it is deemed necessary, the examination will include a rectal prostate examination, but this is seldom needed.



    Routine sperm analysis
    Other
    Biochemical sperm markers
    Capacitation test
    Electron microscopy
    FISH-analysis
    HOS-test
    Survival test
    MAR-test

    A number of tests are designed to determine the quality of your sperm and sperm production. They are all based on the analysis of a sperm sample and all take place in the Andrology lab.

    You can produce a sperm sample at the CRG (in a designated room set aside for this purpose) or do it at home and bring it with you (see practical information). In this case, it is crucial for the test results that you ensure the arrival of your sperm sample in the centre within an hour of its production and that it is kept at body temperature in the meantime. It is also important you catch the whole of the ejaculate. If that is not the case, you must inform us.
    The results of your sperm analysis will be discussed with you by your doctor at the consultation.

    Routine sperm analysis    

    This test is routine and is performed for every man who comes to the CRG for fertility treatment.
    It is carried out according to strict methodology of the WHO, which dictates standard methods relating to the collection of the sample, the delivery thereof to the laboratory and the ensuing laboratory procedures.

    See also sperm quality for the parameters with which the sperm sample must comply, to be regarded as fertile.

    What does a routine sperm analysis determine?
    • A smear evaluates the form and look of the sperm.
    • The vitality of the sperm is also checked: which percentage of sperm is alive in the sample? Not all immobile sperm are necessarily dead.
    • The volume of the ejaculate is also ejaculate, 2 millilitres is the minimum.
    • The acidity (pH) of the sample is measured and the consistency (viscosity) is evaluated.
    • The number of white blood cells is determined (peroxidase test): too many may be an indication of infection. In this case the semen will be placed in culture to determine the nature of infection with a view to treating it.

    Other tests

    Biochemical sperm markers (fructose and alpha-glucosidase)     

    This test is used in azoospermia cases to try to establish why the ejaculate contains no sperm. This does not always mean that none have been produced or ripened; they could be obstructed somewhere around the seminal vesicle and prostate gland, where the seminal fluid is made; or on their way there, in the epididymis or the vas deferentia.
    The test is scientifically based upon the fact that semen is enriched with biochemical and nourishing substances: alpha-glucosidase from the epididymis; fructose when passing the seminal vesicle. By determining which of these substances are present in the seminal fluid, it is possible to diagnose where the blockage has occurred.

    Capacitation test     
    During fertility treatment, sperm is examined to make use of only the highest quality and most motile sperms available in the man’s sample.

    Electron microscopy    
    Sometimes it is necessary to perform an examination under an electron microscope to exclude a particular phenomenon. For instance, in the case where a sperm sample contains exclusively immobile, but nevertheless live sperm. This could be an indication of the (very rare) 'Carthagener Syndrome' in which the tails on the sperm are paralysed. Only an electron microscope can visualise the unimaginably small structures responsible for movement.

    FISH-analysis of sperm    
    This highly specialised test (FISH stands for fluorescent in-situ hybridisation) allows a chromosomal analysis of sperm cells to be made. By way of sample about 3 chromosomes are subjected to meticulous analysis. It is performed in the context of the examination and under certain conditions, for example if there is a large occurrence of abnormal embryos resulting from IVF in a couple having fertility treatment.

    Hypo-osmotic swelling test (HOS-TEST)     
    The HOS-test is a specialised vitality test which establishes whether immobile cells in a sample are dead or alive. It is performed when a sample contains an unusually high number of immobile sperm, sometimes based also on clinical indications. Sperm are placed in a preparation which contains few molecules and is coloured red. This causes the live sperm to swell and shrink. The preparation with eosin (a red disinfectant) allows you to see which ones take up the fluid and which ones don't. The latter ones are the vital, live sperm.

    Sperm survival test   
    This test, which is a more detailed version of the one performed as part of the routine analysis test, determines the vitality of the sperm. The sperm are placed in a medium to see how long they stay alive.

    MAR-test    
    The Mixed Anti-globulin Reaction test is in principle part of the routine analysis test, but is not always included in the test. It determines whether the man produces antibodies against his own sperm (see immunological screening).
    Every hormonal analysis requires a blood sample (see practical information). However, not every blood sample serves this purpose only. Sometimes a blood sample is for immunological or genetic tests

    Hormonal profiling

    Hormonal profiling is standard at your first consultation at the CRG, provided that it has not already been done elsewhere. This test is designed to give a good indication of the sperm production of the testicle.
    More specifically, the levels of FSH hormone are measured, which is the barometer for the function of the testicles:
    • high FSH levels often means a poor function. The sex organs receive the signal from the hypophysis that their production of testosterone, the male hormone which is also secreted by sperm, is optimal.
    • low levels of FSH are equally undesirable, because this leads to too weak a signal for the production of testosterone.
    In some cases, levels of prolactine are investigated. This 'stress hormone' can lead to erection problems in men.


    Basic genetics
    Indications
    Types of tests

    A blood or semen sample are required for genetic screening (see practical information).

    The blood or sperm sample is sent for analysis to the Centre for Medical Genetics (CMG) in UZ Brussel. The CMG provides us with the results, which we then share with our patients at the consultation.

    Basic genetics        

    Important to know

    Our reproductive cells behave differently to other cells in the body:

    • the latter are created by normal cell division or 'mitosis', whereas
    • reproductive cells undergo a complex reduction process during which they retain half of the chromosomes from other body cells.
      During this process certain divisional abnormalities can only be detected in the reproductive cells.

    The human body is made up of many different kinds of tissue, which together form every component of our being. These tissues are made up of individual cells, all of which have a specific function within that tissue. To be able to function properly each cell has a set of on board instructions. This 'recipe book' is in the form of 'chromosomes', receptors which contain genes. In other words, chromosomes are the carriers of our genetic information and are made of DNA.
    Most healthy people have 46 chromosomes, or 23 pairs. There are 22 'normal' pairs and one pair of so-called sex chromosomes. The normal pairs are sorted according to size and numbered 1 to 22. The 23rd pair, the sex chromosomes, is XY in males and XX in females.

    Indications         

    Genetic screening used to be standard in the blood analysis of patients. However, experience has taught us this is unnecessary in the majority of cases. That is why genetic tests are only performed if there is a history of hereditary defects in your family, or indeed in case of specific indications:
    • specifically for men, a low sperm count in the ejaculate can be a reason to perform a genetic test. In cases where there are less than 5 million sperm present in every ml of ejaculate, a genetic cause will be found in 1 in 20 of the cases;
    • all sperm donors are also genetically screened. In any case your Karyotype is determined;
    • genetic screening will be performed on both partners in the following situations:

    Types of tests      

    There are two specific types of genetic tests: cytogenetic (which examines the chromosome in its entirety) and molecular (concentrates on the DNA from which the chromosome is constructed).

    Cytogenetic test    

    To perform this test, white blood cells (lymphocites) are placed in a culture medium. At a pre-determined stage of mitosis - hence the name cytogenetic - the genetic carriers of the lymphocites condense into visual structures, the chromosomes: At this moment we can see how many chromosomes there are and whether they appear normal.

    Karyotype screening

    A cytogenetic test enables Karyotype screening to be performed. Karyotype literally means the character of the chromosomes: amount, length and genetic content. The ingredients of your genetic identity card, so to speak.
    • The test may be performed on men with a strongly reduced sperm count (less than 5 million sperm per ml of semen) to try to find the reason for this. One possible genetic cause could be the Klinefelter syndrome.
    • It is also a standard test for sperm donors. The analysis of the blood sample not only determines the blood type and rhesus factor, but also maps the external characteristics: eye and hair colour, skin type, build, etc.

    When using donor sperm, we always ensure the donor is as similar to the couple as possible regarding blood group and appearance.

    FISH-analysis

    As previously discussed, our reproductive cells behave differently to other cells in the body. During the complex reduction division process, specific ' divisional abnormalities' may occur.
    This is why a specialised test was developed - FISH stands for fluorescence in-situ hybridisation - a process which enables a restricted chromosome-analysis of sperm: Three chromosomes are examined. However, the test is not often performed and only for certain indications, such as when an abnormal number of abnormal embryos develop as a result of IVF in a couple.

    DNA screening      

    Molecular testing goes slightly deeper, i.e. into the DNA structure of the chromosomes to determine which chromosomes display a mutation or deletion of genes. The question here is: where is the receptor fault or where is information missing? This sort of investigation is only restricted to a small number of chromosomes and genes.

    CFTR-gen (mucoviscidosis gene)

    One Belgian out of twenty is a carrier of the mucoviscidosis or CFTR-gene (Cystic Fibrosis Transmembrane Regulator gene). If both partners are carriers, their baby will stand a 25% chance of developing Cystic Fibrosis, a serious lung disease where abnormally large amounts of mucous are produced
    This DNA test is therefore performed on men who have a family member with Cystic Fibrosis and men with azoospermia, due to an absence of the vas deferens at birth. 80% of these men will also be carriers of the faulty mucoviscidosis gene.

    Yq-microdeletion test

    The purpose of this test is also to determine whether a low sperm count (less than 5 million per ml of semen) is due to a genetic defect.
    There are two types of receptors on the Y-chromosome: one makes a man a man, whilst the other on the q-arm of the chromosome, is concerned with the regulation of sperm production. 5% of men with low sperm production have one or more receptors missing from the q-arm of the Y-chromosome (hence the name of the test). A son born to one of these men would also inherit the same genetic problem.

    Routine blood test
    Antibodies against sperm

    Immunological screening of men is based on either a blood sample (see practical information), or a sperm sample (see practical information).

    Routine blood test      

    During your initial consultation at the CRG, and in every fertility treatment case, a blood sample will be taken. The same applies if you wish to become a sperm donor.
    The blood sample is to detect any infections which might be present. Tests will be performed to detect the presence of antibodies against the aids virus (HIV), jaundice (Hepatitis B) and Hepatitis C, and syphilis. The presence of antibodies in the blood indicates that this person is, or has at some time been, infected with the disease in question or that they have been vaccinated against it.
    With certain indications, it is possible to determine whether or not you are infected with chlamydia, gonorrhoea, or other sexually transmitted diseases (STDs). The greatest risk is from chlamydia, because this is the STD most common in the Benelux countries in men and women between the ages of 15 and 35. Women do not always realise that they are infected. In men a chlamydia infection is usually easier to identify, one to three weeks after infection, as pain or a burning sensation during urination and discharge from the penis.
    Chlamydia en gonorrhoea are treatable using antibiotics if the diagnosis is made on time. If you were to infect your partner and she is not treated, it could have serious consequences for her fertility.

    Antibodies against sperm      

    Some men produce antibodies against their own sperm. The following two tests establish whether this is the case. Sometimes both are required to obtain conclusive results.

    MAR-test (Mixed Anti-globulin Reaction test)      
    The MAR-test is performed on a sample with sufficient motile sperm to detect the presence of antibodies. The antibodies would cause the sperm to adhere to each another and render them immotile.
    The other test, the TAT-test is performed using a blood sample and is less precise.

    TAT-test (Tray-Agglutination test)      
    Like the MAR-test, this is performed to detect the presence of antibodies against sperm, but this time in the blood. The same test is also performed on women.
    The test is standard if the man requests a reversal of vasectomy operation.
    This procedure only has a 50% chance of success in restoring fertility, even if it is physically successful. This is because the body becomes aware of the fact that sperm cells, which continue to be produced even after vasectomy, are not going anywhere. In an attempt to dispose of them, antibodies can be produced against them.
    The TAT-test is not flawless and does not provide 100% accurate results regarding the presence of antibodies. It regularly produces false positives: in almost 30% of cases the man appears to be fertile again following a vasectomy reversal, even when the TAT-test suggests otherwise.
    This is why the results are double checked by performing an additional MAR-test, which is more accurate.
    However, a first screening usually involves a TAT-test, because it produces few false negatives. If according to the TAT-test a man does not produce antibodies against his sperm, this is usually not the case either. And the vasectomy reversal can be performed with a greater chance of success.
    Finding the real cause of infertility is rather like doing a jigsaw puzzle. The doctors have to fit all the pieces together by gathering all the results from sperm analysis, physical examination and hormonal profiling for analysis. And still, the answer isn't always obvious.
    Medical image forming techniques can help to solve the puzzle. These examinations do not take place in the CRG itself, but at the Urology of the UZ Brussel. See practical information.

    Scrotal ultrasound

    This is sometimes performed on men whose testicles do not descend at the usual time.
    Using a special ultrasound probe, an image is produced of the testicles, which identifies possible calcification or other abnormalities.
    It can also be done using a duplex doppler to diagnose varicose veins.

    Transrectal ultrasound

    This examination is sometimes performed in cases of azoospermia (the absence of sperm cells in the ejaculate).
    Sometimes results of other tests show a blockage in or around the prostate or the seminal vesicles. This will be the case if the hormonal profile is normal (i.e. no hormonal reason for the azoospermia), whereas the biochemical semen analysis indicates that certain nourishing substances which are normally added to the sperm in the prostate and seminal vesicles are missing in the seminal fluid.
    To perform the scan, a fine probe is inserted into the rectum, with which an ultrasound image of the prostate is generated. It is then possible to determine whether the prostate shows any degree of calcification, or if there are any obvious blockages.

    Practical - How, what and where?

    • The examinations are performed at the Urology department of UZ Brussel.
    • Your doctor at the CRG refers you to a urologist.
    • You need to bring your electronic identity card and your doctor's letter of referral.
    • Register at the Consultation desk, UZ Brussel, entrance C, route 744.
    • At the agreed time you go to the Urology department, entrance C, route 80.
    • Both examinations are outpatient and
      • either without anaesthetic (transrectal ultrasound),
      • or under local anaesthetic (scrotal ultrasound).
    • The results of the examination are given to your doctor at the CRG who will discuss them with you.


    Sometimes, the results of various tests contradict one another. In this case a small surgical procedure may be the only way to obtain a conclusive answer to a particular problem. The aim of a testicular biopsy or aspiration is to determine the number of (healthy) sperm in the testicles. This is where sperm is produced and stays until it is transported to the epididymis to ripen.

    Anatomical explanation     

    Sperm is made in the testicles, inside tiny coiled tubules. The tubules are 30-150ml long and are made of two types of cells: sperm producing cells and feeding cells (Sertolice cells).
    The sperm producing cells are responsible for sperm production. Between the tubules, we find the Leydig cells, which together with the two other types of cell produce male hormones.
    In adult men, the testicle consists of 32% sperm producing cells, 17% Sertoli cell and 3% Leydig cells. The rest is made of supporting tissue.
    The Sertoli cells perform several important functions to do with the production and protection of sperm. They consist of a closed layer, whereby they form a protective barrier between the tubule and the surrounding environment. The layer contains nerve tissue, lymph nodes and blood vessels. It is important that sperm cells are not secreted into the blood. Sperm (production of which starts at puberty) undergoes so many changes in its development, that it becomes alien to the body. The blood-testicle barrier stops rejection from one's own body.
    Other functions of the Sertoli cells include the support, protection and feeding of developing sperm. They also produce a fluid which facilitates the sperm's passage to the epididymis. They also fulfil one other important function: the production of testosterone.

    Causes of azoospermia      

    Azoospermia is the absence of sperm in the ejaculate. This can be caused by an obstruction somewhere along the way to the outside, but can also be a problem in the testicle itself. A testicular biopsy (see below) can show us what is happening in the testicular tubules:
    • bin some men, the sperm producing cells are missing and only the sertolice cells are present (Sertoli cell only syndrome);
    • in others, the sperm producing cells fail to complete the maturation process: maturation arrest
    • sometimes the whole process is carried out, but very few cells are produced (hypospermatogenesis); and
    • the whole tissue of the testicle can be damaged either by infection or trauma (sclerosis or atrophy).
    As a result of all this, no sperm can be found in the sperm sample
    Fortunately, sperm is present in the testicles of about 50% of these men, because normal tubules can often be found next to the empty or damaged ones.

    Testicular biopsy       

    Under local or general anaesthetic, tiny amounts of testicular tissue are removed and sent for examination to the pathologist.
    The type of anaesthetic used depends on your personal choice and other medical factors. If more biopsies need to be removed, a general anaesthetic is recommended.

    The biopsies will show whether the testicles have (sufficient) functional tubules and whether sperm production is normal.

    In the case of azoospermia, the operation will provide conclusive results as to whether the absence of sperm in the ejaculate is due to a blockage:
    • a production fault in the testicles (testicular cause), or
    • or a blockage further up (post-testicular).
      In the latter case the biopsy will contain sufficient amounts of sperm.

    The operation is not only performed for examination purposes: at the same time we can look for sperm on other tissue (see TESE) and, if any are found, they can be immediately frozen for later use.
    In approximately half of all men with an abnormal sperm production, sperm can be harvested from the tissue, because normal tubules exist among the name abnormal ones. Sometimes taking several pieces of tissue is necessary to find a needle in a haystack.