Chapter 5a
Beyond the Semen Analysis

from the book How to Have a Baby: Overcoming Infertility
by Dr. Aniruddha Malpani, MD and Dr. Anjali Malpani, MD.

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For the man with a poor semen sample, additional tests which may recommended include specialized sperm tests; blood tests; and testis biopsy.

Antisperm Antibodies Test

The role of antisperm antibodies in causing male infertility is controversial, since no one is sure how common or how serious this problem is. However, some men (or their wives) will possess antibodies against the sperm, which immobilize or kill them and prevent them from swimming up towards the egg. The presence of these antibodies can be tested in the blood of both partners, in the cervical mucus, and in the seminal fluid. However, there is little correlation between circulating antibodies (in the blood) and sperm-bound antibodies (in the semen), There are many methods of performing this test, which can be quite difficult to standardize, as a result of which there is a lot of variability between the result reports of different laboratories. The older methods of testing used agglutination methods on slides and in test tubes. Perhaps, the best method available today is one such uses immunobeads, which allow determination of the location of the antibodies on the sperm surface. If they are present on the sperm head they can interfere with the sperm’s ability to penetrate the egg; if they are present on the tail they can retard sperm motility. Of course, if the test is negative, this is reassuring; the problem really arises when the test is positive! What this signifies and what to do about it are highly vexatious issues in medicine today, and doctors are even more confused about this aspect than the patients.

Semen Culture Test

Ion the semen culture test, the semen sample is tested for the presence of bacteria, and , if present, their sensitivity to antibiotics is determined. Interpreting this test can also be problematic! It is normal to find some bacterial in normal semen samples - and the question which must be answered is : are these bacteria disease- causing or not?

Tests which assess the sperm’s ability " to perform" include the following sperm function tests.

Postcoital Test (PCT)

The postcoital test is the easiest test of sperm function, since it is performed in vivo. It is done when the wife is in the " fertile" period, during which time the cervical mucus is profuse and clear. The gynecologist examines a small sample of the cervical mucus, under the microscope, a few hours after intercourse. ( This can be embarrassing and awkward for the patient, but it is not painful at all). Finding 5-10 motile sperm per high power microscopic field means that the test is normal. A normal test implies normal sperm function and can be very reassuring.

An abnormal test needs to be repeated and, if the problem is persistent, one needs to determine if the defect lies in the sperm or in the mucus, by cross-testing with the husband’s sperm, donor sperm, wife’s mucus and donor mucus.

Bovine Cervical Mucus Test

The bovine cervical mucus test is another form of testing for the ability of the sperm to penetrate and swim through cervical mucus, with the difference that in this case, the mucus used is that of a cow (since this is commercially available abroad in a test kit.) The sperm are placed in a column of cervical mucus and how far the sperm can swim forward through the column in a given amount of time is checked with the help of a microscope.

Sperm Viability or Sperm Survival Test

This is a simple test, which provides crude (but useful!) information on the functional potential of the sperm. The sperm are washed using the same method which is used for IVF (either a Percoll spin or sperm swim up) and the washed sperm are then kept in a culture medium in the laboratory incubator for 24 hours. After 24 hours, the sperm are checked under the microscope. If the sperm are still swimming actively, this means that they have the ability to "survive" in vitro for this period- and this is reassuring. If, however, none of the sperm are alive after 24 hours, this suggest that they may be functionally incompetent.

Sperm Penetration Assay (SPA, Hamster Assay)

Since the basic function of a sperm is to fertilize an egg, scientists were very excited when they found that normal sperm could penetrate a denuded (zona-free) hamster egg. A zona-free hamster egg is obtained from hamsters egg. A zona-free hamster egg is obtained from hamsters and the covering (the zone) removed by using special chemicals. The egg are then incubated with the sperm in an incubator in the laboratory. After 24 hours, the eggs are checked to ascertain how many sperm have been able to penetrate the egg. The result gives a penetration score, which gives an index of the sperm’s fertilizing potential. This is a very delicate technique and is not available in India. In any case, nowadays scientists the world over are quite disenchanted with the test, since the correlation between IVF results (the ability to fertilize human eggs) and the SPA (the ability to penetrate zona-free hamster eggs) is quite poor.

• Testing for acrosomal status
• HOS test - hypo-osmotic swelling test-which tests for the integrity of the sperm membrane
• CASA - computer-assisted sperm analysis
• Hemizona assay
• Electron microscopy of sperm

The aforementioned tests are highly sophisticated and are not easily available. Another drawback is that these tests are often not standardized adequately, so that interpreting their results can be quite difficult.

The ultimate sperm function test is the IVF, since this directly assesses whether or not the husbands" sperm can fertilize the wife’s eggs. The best way to perform this test is to culture some of the eggs with the husband’s sperm and the others with donor sperm of proven fertility, at the same time. If the donor sperm can fertilize the eggs, and the husband’s sperm fail to do so, then the diagnosis of sperm inability to fertilize the egg is confirmed. However, even this test is not infallible, since it has been shown that about 5% of sperm samples which fail to fertilize an egg in the first IVF attempt, can do so in a second attempt at IVF.

by Dr. Aniruddha Malpani, MD and Dr. Anjali Malpani, MD.

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