1. Semen Analysis

2. Objective: Physiology of semen formation Indication of semen analysis
Macroscopic examinationof semen
Microscopic examination
Antibody testing in semen
Reference ranges

3. Physiology of Seminal fluid
Semen is grey opalescent fluid , which consists of suspension of spermatozoa in seminal plasma.
It is made up of the secretions of all accessory glands of the male genital tract.

4. The Percentage contribution of Each of the Secretions in Seminal Fluid
% Ejaculate
Source of Secretion 5
Testis
40-80
Seminal vesicles
13-33
Prostate
2-5
Bulbo-uretheral & uretheral glands

5. Fructose: imp energy for sperms especially needed for sperms motility
Prostaglandins: play a role in controlling sperm movement & sperm penetration of cervical
mucus
Fibrinogen-like substance : acted upon by enzyme vesiculase to induce clotting that occurs in semen
The prostatic fluid pH =6.5
Enzymes: vesiculase
Acid phosphatase

6. Bulbo-uretheral & uretheral glands
The fluid rich in mucoprotein , so it lubricates the urethra
Occasionally in the secretions contain antisperm antibodies( may be of importance in infertility cases)

8. During ejaculation , each of the components that make up semen may be discharged from the urethra in a sequence
The first part is made up of testicular component of semen followed by secretion of the prostate , lastly the secretion of the vesicles are expelled.

9. Indication of Semen Analysis
Assessment of fertility/infertility ( most common)
Determination the effectiveness of vasectomy
Determination of suitability of semen for artificial insemination
Follow up of fertility after cancer treatment by radio- or chemotherapy
Forensic purpose

10. Semen analysis Specimens:
2 samples should be collected for initial evaluation,
the interval between the two collections should not be less than 7 days or more than 3 weeks ,
if the results of these assessments are markedly different , additional samples should be examined

11. Examination of Semen Macroscopic Examination Microscopic Examination
Volume
Liquefaction time
Appearance pH
Viscosity
Microscopic Examination
Sperm count
Sperm motility
Sperm viability
Sperm morphology
Agglutination
Antibodies coating of sperms
Biochemical Tests
Fructose test
Acid phosphatase

12. Macroscopic Examination

13. Volume : ( 2-6ml/ ejaculation)
Aspermia: total absence of ejaculation ( rare)
Hypospermia or oligospermia: the seminal fluid is < 2ml
Hyperspermia: volume > 10 ml
( spermia denotes seminal fluid not spermatozoa)

14. Liquefaction time:
( forms gel-like clot immediately after ejaculation)
a normal semen sample liquefies with mint .
( prolongation must be recorded).

15. Appearance: homogeneous , grey-opalescent appearance
The color is due to high content of protein &presence of > 60 million sperms/ml
May be less opaque if sperm concentration is very low
Dense white turbid in inflammation and high WBCs
Haematospermia :when red blood cells are present (blood)
Yellow in patient with jaundice (very bright yellow due to bilirubin) or taking some vitamins
Or contaminated with urine ( uriniferous odour)

16. pH : between7.2-8.0 recorded in fresh semen by using pH paper .
A patient exceeding pH 8.0 may suggest acute disease of the seminal vesicles
Lowering of pH may be due chronic inflammatory of seminal vesicles
Or contamination with urine

17. Viscosity :
Normal viscosity is that which allows semen to be poured drop by drop out of the container.
In case of abnormal viscosity drop will form a thread more than 2 cm long
High viscosity : can interfere determination of sperm motility

18. Microscopical Examination

19. Sperm Count: Total number of sperms in an ejaculation.
It is 20 million/ml i.e. 60 million / ejac
It is obtained by multiplying the sperm concentration by the volume
Azoospermia: means no spermatocytes ( male sterility)
Oligozoospermia: < 20 million/ml (less than 50 million/ejaculation
Polyzoospermia : may reach 350millions/ejaculation

20. Sperm Count Decreased: vasectomy varicocele
primary testicular failure (Klinefelters)
secondary testicular failure
congenital vas deferens obstruction
endocrine causes (prolactinemia, low testosterone)

21. Assessment of sperm motility:
the motility of each spermatozoon is graded :
‘a’ =rapid progressive motility
‘b’= slow or sluggish motility
‘c’= non progressive motility
‘d’= immotility

22. Viability should be determined if the % of
immotile spermatozoa exceeds 50 %
Supravital stain:
Eosin /Nigrosin
Viable do not take up the stain

23. Assessment of sperm morphology

24. Normally the sperm count contains fewer than 20 % abnormal forms e. g
Normally the sperm count contains fewer than 20 % abnormal forms e.g. bitailed, short tailed , 2 heads …..etc.

25. Other cellular elements:
Leukocytes:
leukocytes predominantly neutrophils are present in most human ejaculate.
Normal <1x106 /ml or < 5/HPF
5-20 WBC/HPF =increased
20-40 WBCs/HPF =+
>40 WBCs/ HPF = ++
If increased leukospermia or pyospermia may be associated with infection and poor sperm quality
When the number is increased, microbiological tests should be performed to investigate the presence of of infection in any of the accessory glands

26. Agglutination
Reported when motile sperm stick to each other in a definite pattern.
Head-head
Tail-tail
Head-tail
Immunological cause of infertility

27. Biochemical Tests Fructose Test: (a) absence of seminal vesicle;
It is secreted for sperm nutrition from seminal vesicle
Impairment of seminal vesicular secretion will result in reduced fructose secretion in semen and the motility of the sperms will be reduced.
Fructose disappears in cases of :
(a) absence of seminal vesicle;
(b) obstruction of ejaculatory duct; and
(c) inflammation of seminal vesicle. It is decreased in case of testosterone deficiency. So, fructose is used as fertility test.

28. Secreted from the prostate. The test is used as:
Acid phosphatase:
Secreted from the prostate.
The test is used as:
1)A marker of prostatic functions;
and
2)In forensic laboratories as a test for the presence of
semen.

29. Testing for Antibody coating of spermatozoa
Sperms can induce immune response not only between species but also within one species and within an individuals
Sperm antibodies may :
Cause agglutination of sperm
Cause reduction in motility
Inhibit the ability of sperm to penetrate cervical mucus
Impede binding of the sperm to the oocyte
Antibodies may be IgA & IgG
It must be remembered that sperm antibodies may be present in the cervical mucus itself , may inhibit the entry of sperm in the female genital system

30. Reference Ranges

31. Total count > 40 million/ejaculate
Volume ml pH
Color: greyish white
Liquefaction: min.
Count >20 million/ml
Total count > 40 million/ejaculate
Motility: > 50 % motile( grades a+b) or > 25% with progressive motility ( grade a) within 60 minutes of ejaculaton
Morphology > 30% normal form
Viability > 75% viable
WBC< 1million/ml
RBC none
Quoted from the WHO Laboratory Manual for the Examination of Human Semen and Sperm-Cervical Mucus Interaction ,1999