Semen examination

Semen is a “ thick, viscous, creamy, slightly yellowish or grayish” substance made up of  spermatozoa — commonly known as sperm — and a fluid called seminal plasma, secret from the male reproductive organs.
The function of seminal plasma are:
To provide motility to sperm
To provide nutrition to spermatozoa

1. SEMEN ANALYSIS Ms Ankita R Bhatiya Assistant Professor SHREE P.M.PATEL COLLEGE OF PARAMEDICAL SCIENCE N TECHNOLOGY

2. SEMEN ANALYSIS It include: 1.What is Semen? 2.Production of Semen. 3.Indication of Semen 4.Collection of Semen. 5. Examination of Semen.

3. SEMEN ANALYSIS 1.What is Semen? Semen is a “ thick, viscous, creamy, slightly yellowish or grayish” substance made up of  spermatozoa — commonly known as sperm — and a fluid called seminal plasma, secret from the male reproductive organs. The function of seminal plasma are: To provide motility to sperm To provide nutrition to spermatozoa

4. SEMEN ANALYSIS 2.Semen production: The male reproductive system includes the penis, scrotum, testes, epididymis , vas deferens, prostate, and seminal vesicles. The penis and the urethra are part of the urinary and reproductive systems. The scrotum, testes (testicles), epididymis , vas deferens, seminal vesicles, and prostate comprise the rest of the reproductive system.

5. SEMEN ANALYSIS Overview of the Male Reproductive System The  penis  consists of the root (which is attached to the lower abdominal structures and pelvic bones), the visible part of the shaft, and the glans penis (the cone-shaped end). The opening of the urethra (the channel that transports semen and urine) is located at the tip of the glans penis. The base of the glans penis is called the corona. In uncircumcised males, the foreskin (prepuce) extends from the corona to cover the glans penis .

6. SEMEN ANALYSIS The  scrotum  is the thick-skinned sac that surrounds and protects the testes. The scrotum also acts as a climate-control system for the testes because they need to be slightly cooler than body temperature for normal sperm development. The cremaster muscles in the wall of the scrotum relax to allow the testes to hang farther from the body to cool or contract to pull the testes closer to the body for warmth or protection. The  testes  are oval bodies that average about 1.5 to 3 inches (4 to 7 centimeters) in length and 2 to 3 teaspoons (20 to 25 milliliters) in volume. Usually the left testis hangs slightly lower than the right one. The testes have two primary functions: The Testes and Ovaries Producing sperm (which carry the man's genes) Producing testosterone (the primary male sex hormone)

7. SEMEN ANALYSIS The  epididymis  consists of a single coiled microscopic tube that measures almost 20 feet (6 meters) in length. The epididymis collects sperm from the testis and provides the environment for sperm to mature and acquire the ability to move through the female reproductive system and fertilize an ovum. One epididymis lies against each testis. The  vas deferens  is a firm tube (the size of a strand of spaghetti) that transports sperm from the epididymis. One such duct travels from each epididymis to the back of the prostate and joins with one of the two seminal vesicles. In the scrotum, other structures, such as muscle fibers, blood vessels, and nerves, also travel along with each vas deferens and together form an intertwined structure, the spermatic cord. The  urethra  serves a dual function in males. This channel is the part of the urinary tract that transports urine from the bladder and the part of the reproductive system through which semen is ejaculated.

8. SEMEN ANALYSIS The  prostate  lies just under the bladder and surrounds the urethra. Walnut-sized in young men, the prostate enlarges with age. When the prostate enlarges too much, it can block urine flow through the urethra and cause bothersome urinary symptoms.  The  seminal vesicles,  located above the prostate, join with the vas deferens to form the ejaculatory ducts, which travel through the prostate. The prostate and the seminal vesicles produce fluid that nourishes the sperm. This fluid provides most of the volume of semen, the fluid in which the sperm is expelled during ejaculation. Other fluid that makes up a very small amount of the semen comes from the vas deferens and from Cowper glands in the urethra.

9. SEMEN ANALYSIS 3.Indications for semen analysis: 1. Investigation of infertility 2. Post - vasectomy by confirming absence of sperm. 3. To support or disprove a denial of paternity on the grounds of sterility. 4. To examine vaginal secretions or clothing stains for the presence of semen in medico legal cases. 5. For selection of donors for artificial insemination. 6. For selection of assisted reproductive technology, e.g. in vitro fertilization, gamete intrafallopian transfer technique. 7. After reversal of vasectomy to confirm the success of procedure.

10. SEMEN ANALYSIS 4.Sample collection - Preparation Sexual abstinence – 2- 7 days Two separate samples at least 7 days apart should be analyzed The duration of abstinence should be constant Sample collection - private room in the same laboratory Recommended procedure – Masturbation Pre warmed (21oC), sterile, non-toxic, a clean, dry,wide mouthed plastic/glass container.

11. SEMEN ANALYSIS 5. Precaution: The sample should not be collected in condoms, since the powder or lubricant applied to the condoms may be spermicidal. The container in which semen sample is collected should be free from detergents. The specimen collected by masturbation in clinical pathology laboratory. This allow a complete examination of the semen particularly the liquefaction time. In the patients house by masturbation, the specimen should be delivered within 30 min to the laboratory.

12. SEMEN ANALYSIS 6.Storage: The semen specimen should be examined immediately after collection. It is necessary to store then do not store in refrigerator, store at room tem.

13. SEMEN ANALYSIS Clinical significance: Low sperm count are observed when there is suppression of gonad tropic hormone estrogen or androgen. Hyper & Hypothyroidism,trauma,infection , damage of testis. Oligozoospermia & azoospermia .

14. SEMEN ANALYSIS Examination of Stool: Physical examination: Chemical examination: Microscopic examination:

15. SEMEN ANALYSIS Physical examination: 1.Volume: Normally: b/w 2.5 to 5ml. Low semen volume causes: – Obstruction of the ejaculatory duct – Congenital bilateral absence of the vas deferens ( CBAVD), a condition in which the seminal vesicles are also poorly developed. – Partial retrograde ejaculation – Androgen deficiency – Inadequate erection & improper mood at collection – Incomplete collection High semen volume – may reflect active exudation in cases of active inflammation of the accessory organs

16. SEMEN ANALYSIS 2. Color: Whitish or grayish Slightly yellow – prolonged abstinence Deep yellow – pyospermia , jaundice or taking certain vitamins or drugs Red-brown - red blood cells are present ( haemospermia ) Trauma to the genital tract Inflammation Tumor of the genital tract .

17. SEMEN ANALYSIS 3.Appreance: Viscid, Opaque Less opaque : if sperm concentration is very low 4.Viscosity: when ejaculated, semen is fairly viscid and it falls drop by drop Viscosity of the sample estimated After liquefaction Procedure : gently aspirate sample into a wide-bore 5ml plastic disposable pipette  allow the semen to drop by gravity. Observe the length of any thread. A normal sample leaves the pipette in small discrete drops. If viscosity is abnormal , the drop will form a thread more than 2 cm long.

18. SEMEN ANALYSIS 5. Liquefaction time: Normal – 15- 30 minutes after collection The complete sample usually liquefies within 15 minutes at room temperature If the semen does not liquefy within 30 minutes, do not proceed with semen analysis but wait for another 30 minutes. Rarely it may take up to 60 minutes or more. If complete liquefaction does not occur within 60 minutes, this should be recorded. Semen samples collected at home or by condom will normally have liquefied by the time they arrive in the laboratory.

19. SEMEN ANALYSIS 2. Chemical examination: 1. FRUCTOSE TEST This test determines androgen deficiency or ejaculatory obstruction of semen ; the level of seminal fructose is low in both these conditions. Normal seminal fructose level is 150-300mg/dl Fructose is measured by qualitatively by resorcinol test.

20. SEMEN ANALYSIS Procedure: 5 ml resorcinol reagent + 0.5 ml of semen in a test tube Mix and place in a boiling water bath for 5min or heat. Observations : Red colored ppt. in 30 seconds. In quantitative assays, this is compared with a known fructose standard at 490nm. Normal level of fructose: 150-300mg/dl. Reduced levels: Seminal vesicle dysfunction, High sperm count, obstructed vas deferens

21. SEMEN ANALYSIS 2.PH: Reference value - ≥ 7.2 Measured after liquefaction , preferably after 30 minutes Should be measured within 1 hour of ejaculation since it is influenced by the loss of CO2 that occurs after production. Procedure: For normal samples: pH paper in the range 6.0 to 10.0 should be used. For viscous samples, the pH of a small aliquot of the semen can be measured using a pH meter designed for measurement of viscous solutions. pH < 7.0 – With absence of sperm - ejaculatory duct obstruction or congenital bilateral absence of the vas deferens, a condition in which seminal vesicles are also poorly developed. – contaminated with urine. Semen pH increases with time, as natural buffering decreases, so high pH values may provide little clinically useful information.

22. SEMEN ANALYSIS 3. Microscopic examination: 1.Sperm Motility: Procedure: Place a drop of liquefied semen on clean glass slide. Put a cover slip over it and examined it under the microscope . Normally , within 2 hours of ejaculation , more than 60% of spermatozoa are vigorously motile and in 6-8 hours 95-60% are still motile is. If motility is less than 50%, a stain for viability such as methylene blue or eosin Y with nigrosin as counte rsatin can be done. Heads of non-motile sperms are stained positive for the dye. Sterile moles have <25% motility of sperms Motile forms decrease by about 5% per hr after the 4 th hr following collection ,motility less than 60% may be associated with infertility.

23. SEMEN ANALYSIS Types of Motility : Darting motility : The sperm are moving on their self.(Grade-4) Very strong activity with forward progression- (Grade 3) Decent sperm activity with forward progression- (Grade 2) Sluggish motility : The sperm are active and are transferred to other place very slowly.(grade-1) Necrozoospermia : Sperm are present but they are immotile they can not move.(Grade-0) Oligozosopermia : Here few sperms are only motile.(Grade-2) Azoospermia : Sperm are present in high power.(Grade-0)

24. SEMEN ANALYSIS 2.Sperm count: Sperm count is same as WBC count. The calculation is similar to WBC formula expect that the reported sperm count is per ml instead of per cumm . So result is multiplied with thousand to convert cumm to ml. The composition of diluting fluid is as under: Sodium bicarbonate 5 gm Formalin (neutral) 1 ml Distilled water 100 ml This is done in Neubauer’s ( hemacytometer ) chamber using a WBC pipette. Draw liquefied semen in WBC pipette up to marks 1 and then the diluting fluid up to mark 11. After mixing it properly, charge the chamber. Allow the spermatozoa to settle down in 2 minutes. Examine under microscope and count the number of spermatozoa in one large peripheral square. Normal Range: 40-300 millions/ml

25. SEMEN ANALYSIS 3.Sperm morphology: Procedure : Prepare a thin smear from liquefied semen on a glass slide and stain with a Romanowsky stains , Observe at least 200 sperms for any abnormality in their morphology Normally 80% of sperm are normal The abnormal forms of sperms are with double head , swollen and pointed head , double tail and rudimentary forms. Observation: 1.Spermatozoa head: Light blue 2.Nuclear posterior: Dark blue 3.Bodies & tail: Red or pink 4.Spermatozoa size: 50-70 UM Observation for other abnormalities : Abnormal:0-20% More than 20% abnormal form may be associated with infertility.

26. SEMEN ANALYSIS Following abnormalities are observed in spermatozoa: