Semen Analysis- The Male Factor

Sperm fertilizes an egg to conceive a baby. The society earlier tended to blame women alone for infertility.  Now technology has proved that men are just as responsible for infertility, if not more.

 “WHO statistics show that the male factor accounts for 51.2 % of infertility cases and the female factor for 48.8% cases. It is well known that men must share equal responsibility,” said Dr. A.K. Bhattacharyya, a retired Biochemistry Professor. 

Semen Analysis video

Sperm development

An adult man produces millions of sperm cells every day. An individual sperm is extremely small in size; it is about 0.05 millimeters long. The sperm develops in the testicles within a system of tiny tubes called the seminiferous tubules. At birth, these tubules contain simple round cells, but during puberty, testosterone and other hormones cause these cells to transform into sperm cells. The cells divide and change until they have a head, mid-section or neck and tail, like tadpoles. This tail helps in swimming. The sperm completes its development in epididymis. It takes about 4 to 6 weeks to travel through the epididymis.

Human sperm

Sperm journey

The sperm then move to the vas deferens, or sperm duct. These seminal vesicles and prostate gland produce a whitish fluid called seminal fluid, which mixes with sperm to form semen. When a male is sexually stimulated the semen is pushed out of the male's body through the urethra called ejaculation. Each time a man ejaculates, it can contain up to 500 million sperms.

When the sperm enters the female reproductive organ, fertilization or conception takes place. If the fertilization does not occur, both the partners are advised to go for a medical check up.

Male factor and the cause

Male infertility can be caused by a number of factors, including problems associated with sperm production, sperm transport, and sperm motility as well as anatomical problems like blockage of the vas deferens

Sperm motility is also affected by temperature. If the testicles are too warm, the sperm motility and possibly the sperm count may be reduced.

In some cases, sexually active men or women may also develop "sperm antibodies" which may impair sperm function. This means that a person becomes "allergic" to sperm.

 “Life style changes force us to move further and further away from nature. We consume food with pesticide and other chemical components. Pesticide-laden food is one of the major causes for lowering of sperm count; today an average man does not produce more than 20 million is the sperms,” said Dr. A.K. Bhattacharyya.

Hormonal changes, medications, alcohol, smoking, testicular injury, stress and exposure to environmental toxins can cause infertility.

Approaching doctor

It is important to provide the doctor with a complete health history of the patient. The use of medications, alcohol, cigarette smoking and exposure to chemicals or heavy metals.

Whether the patient had diseases like mumps, chicken pox, measles, TB, jaundice or any problems associated with sex and so on.

“When we get the result of the analysis particularly the functional properties we try to co-relate with the finding of the history which gives a very good clue,” said Dr. A.K. Bhattacharyya.

Obtaining semen sample

      When a patient is asked to collect the semen sample for examination these are to keep in mind.

1.      Sexual abstinence for at least 2 days before obtaining the sample.

2.      Obtain the sample by manual masturbation only. No vaginal, oral or anal contact with the penis. Avoid using lubricants.

3.      Collect the entire semen specimen in a clean, wide mouthed, plastic or glass container.

4.      Label the specimen with names, date, and time of collection.

5.      The specimen must arrive at the lab without delay.

Examination

Examination of the semen involves determination of the semen amount, sperm morphology, and sperm motility.

      Liquefaction test

The semen sample is kept in incubator for at least 30 minutes in order to liquefy. The incubator is maintained at 37⁰ C. Normally semen is produced as a coagulum. When semen falls drop by drop it is said to be normal. If semen does not trickle in drops it is abnormal. 

 

Semen amount test

According to the World Health Organization, the normal semen volume is 2-5 ml.

      

      Sperm Morphology test

It deals with the size, shape and appearance of the sperm. The characteristics should be assessed by carefully observing a stained sperm sample under the microscope.

                   

      Semen analysis process

The semen is taken out and dropped on a slide. It is smeared and fixed with ethanol ether and kept for 5 to 10 minutes. Then the sample is dipped in different levels of alcohol, starting from 80%, 70%, 50% till it reaches distilled water. This process is called hydration. 

Now it has to be stained in haematoxylin stain. After keeping it for 3 minutes rinse in tap water.

Now reverse the process. Dip the slide in 50 %, 70%, 80% and 90% alcohol. This is called dehydration.

For better staining the sample has to be dipped in Orange G for 3 minutes. After dipping in 90% alcohol stain the sample in EA 50. Again dip it in 90% of alcohol followed by Xylene.

The sample is taken to a microscope and adjusted at 100x magnification. An emersion oil is applied for perfect assessment.

Several shapes or forms have been identified and characterized. They fall into normal forms, abnormal head, abnormal neck, and abnormal tail.

                                                   Normal and abnormal sperm

      Sperm Motility test

Sperm motility is the study of moving sperm in an ejaculated specimen.

Specific quantity of sample is taken and with the help of the microscope the sample is analyzed. Usually 20x magnification is used for counting purpose.

The sperm motility is classified into rapid progressive, slow progressive, non progressive and immobile.

The sperm must show more than 50% of rapid progressive and slow progressive to achieve fertilization.  This is rapid progressive, this is slow progressive, this is non progressive and this is immobile

DNA integrity test

DNA integrity test is a better diagnostic and prognostic marker of sperm reproductive potential. Sperm DNA damage has been closely related with numerous indicators of reproductive health including fertilization, implantation, embryo quality, spontaneous abortion, congenital malformation and childhood diseases.

The sample is Smeared on a slide and fixed it on carnoy’s fluid. The role is to fix and to enhance the DNA damage. It allows the acridine molecules to react with the sperm. The sample should be kept overnight and washed in water. Let the sample dry on a tissue paper. Immerse the slide in acridine orange solution and wash it in water. Keep the slide on a place where there is no light.

The green color is normal and the red color is abnormal.

                       Human Sperm DNA integrity Acridine Orange test

Semen acid-base balance test (pH) 

The pH of semen is measured by using a special paper blot that changes color according to the pH of the specimen. Prostatic secretions are acidic while the secretions of the seminal vesicles are alkaline. Therefore, alterations in pH may reflect a dysfunction of one or both of these accessory glands.

“The normal pH count ought to be between 7.6 and 8. If the count goes down, the condition is called high prostatic activity. In other words, it causes prostatic problem.  If the count is higher, the cell may die, but there will be no prostrate problem in the future. So pH count measures the function of of two organs: prostate and seminal vesicle,” said Dr. A.K. Bhattacharyya.

Gradient methodology

"You can pick out the best cell without any problem by a particular methodology called gradient methodology. These media are not available in India; we have to import them from abroad. Through the media by gradient procedure we can separate the normal cell from the abnormal cell. You must remember that only normal cells can be used for Assisted Reproduction Technology, IUI, IVF or ICSI. The choice of a particular technology depends on the number of cells available after separation. If the number of available normal cells is high, IUI is the preferred option. If the cell count is relatively low, IVF is possibly the best option. If the count is very low, you have to go for ICSI,” said Dr. A.K. Bhattacharyya.

Ray of hope

After all those tests, if the semen does not contains sufficient healthy sperms, there are at least three other possibilities which will give a ray of hope to infertile couple.

The best sperm is selected and treated with the egg for fertilization through the process called Intrauterine Insemination (IUI), In Vitro Fertilization (IVF), Intra Cytoplasmic Sperm Injection (ICSI) and so on depending on the individual

Intrauterine Insemination (IUI): Semen is processed in the laboratory to enhance the proportion of normal active sperm. It is then placed directly into the uterus using one of many various specialized catheters.

In Vitro Fertilization (IVF) or test tube baby: Eggs are obtained from the ovaries and cleaned. Processed sperm is then added to the droplet containing the egg. Fertilization then occurs. After three to five days the embryos are placed in the uterus.

Intra Cytoplasmic Sperm Injection (ICSI): A process where a single sperm is injected into an oocyte (egg or ovum that is produced inside the ovary) using micromanipulation equipment. Useful when there are very few sperms.

You can watch my documentary: Semen Analysis

Courtesy EMMRC Kolkata