2. OUTLINE
• What is semen
• Formation of the sperm cell
• WHY PERFORM SEMEN ANALYSIS
• What is the “standard” approach to semen
evaluation?
Hassaan Ali 20154/7/2015
3. What is semen
A mixture of seminal plasma and cells
• Seminal plasma contains:
– Prostatic fluid (~30% of the volume)
– Epididymal plasma (~5% of the volume)
– Seminal vesicle fluid (the remainder of the ejaculate)
• The cells are:
– Spermatozoa
– Germ line cells
– Leukocytes of various types
– Bacteria
– Epithelial cells
– Occasional red cells
Hassaan Ali 20154/7/2015
6. Spermatogenesis is a cascade of cell divisions:
Mitosis: spermatogonia to
primary spermatocytes
First meiotic division:
secondary spermatocytes
Second meiotic division:
haploid spermatids
This process takes 70 ± 4 days
in the human – so errors will
take about 3 months to show
up
Hassaan Ali 20154/7/2015
9. WHY PERFORM SEMEN ANALYSIS?
•Diagnosis of sterility
•Diagnosis of infertility
•Prognosis for fertility
•Identify treatment options:
• surgical treatment
• medical treatment
• assisted conception treatment
Hassaan Ali 20154/7/2015
10. What is the “standard” approach to
semen evaluation?
•World Health Organization’s Lab Manual fifth edition in 2010..
•Focus is on standardization with expanded section on quality
control.
•Methods amenable for use in any (“third world”) country.
•Basic infertility work-up.
Hassaan Ali 20154/7/2015
11. Sample Collection
• For a meaningful result, semen samples must always be
collected under standardized conditions:
– the container has to be sterile and known NOT to be
spermotoxic (i.e. provided by the lab)
– the man must have had 3 – 5 days of abstinence
– the man must have washed his hands before collection
(particularly if microbiological analysis is requested)
– the sample must be kept at 37°C until analysis, which begins
ideally within 30 min, but absolutely within 60 min, of
ejaculation
Hassaan Ali 20154/7/2015
12. Methods Collection
1. Masturbation (the method of choice for all seminal fluid
tests).
2. By condom: it is not recommended for fertility testing
because the condoms may contain spermicidal agents.
3. By coitus interrupts: (withdrawal method).
4. Percutaneous epididymal sperm aspiration (PESA).
if a blockage in the vas deferens is suspected, semen can be
taken directly from the epididymis
5. TESA: Testicular sperm extraction (TESE)- Open Testicular
Biopsy: is a highly invasive, open surgical procedure
performed under general anaesthetic. The scrotum and
testes are cut open, before testicular tissues are cut away
and examined for sperm, which, if present can be extracted.
Hassaan Ali 20154/7/2015
13. Semen Study
– There are several macroscopic and microscopic evaluations
which give useful diagnostic information about the sample
–Macroscopic microscopic
– Appearance
– Odour
– Liquefaction
– Volume
– Viscosity
– pH
-concentration/count
- motility
-morphology
-viability
Hassaan Ali 20154/7/2015
15. Nomenclature for semen variable
(WHO)
• Normal semen quality Normospermic
• No ejaculate Aspermia
• Low volume Hypospermia
• High volume Hyperspermia
• No spermatozoa Azoospermia
Hassaan Ali 20154/7/2015
16. Nomenclature for semen variable
(WHO)
• Low spermatozoa conc. Oligozoospermia
• Low spermatozoal motility Asthenozoospermia
• Low normal morphology Teratozoospermia
• Excessively high sperm conc., Polyzoospermia
• WBCs in semen Pyospermia
• RBCs in semen Heamatospermia
• Non viable sperms (dead). Necrozospermia
Hassaan Ali 20154/7/2015
17. Factors affecting SA results
• Medicines senomroh elamef dna elam ,enidtiemic sa hcus ,
dna ,niotnarufortin ,enizalasaflus ,)negortse,enoretsotset(
senicidem yparehtomehc emos.
• Caffeine occabot gnikoms dna ,anaujiram ,eniacoc ,lohocla ,.
• Herbal fo sesod hgih dna trow s'nhoJ .tS sa hcus ,senicidem
aecanihce.
• Temperature -fi wol yletaruccani eb lliw eulav ytiltiom mreps
dloc steg elpmas nemes eht.
• Exposure to radiation niatrec sa hcus( slacimehc emos ,
erusopxe taeh degnolorp dna ,)sedicimreps ro sedictisep.
• An incomplete semen sample --si elpmas a fi nommoc erom
notiabrutsam naht rehto sdohtem yb detcelloc.
• Not ejaculating for several days -- affect the semen volume
Hassaan Ali 20154/7/2015
18. Macroscopic
• Volume
• Normal:1.5 ml per ejaculation
• Low volume (<1ml) reflect a problem with the seminal vesicles and
prostate – a block, retrograde ejaculation, infection or lack of
androgen.
• Low semen volume cannot neutralize vaginal acidity
• High semen volume dilute sperms/ active infection
• pH
• Normal:=/>7.2 (alkaline)
• Increased pH is indicative of infection within the reproductive tract.
• A decreased pH Seminal vesicle dysfunction or agenesis.
• Vas deferens agenesis.
• Ejaculatory ducts obstruction.
• Incomplete collection.
• .
Hassaan Ali 20154/7/2015
19. Macroscopic
• Appearance
• Normal:Whitish to grey opalescent
• Yellow (urine, jaundice);
• Pink/Reddish/Brown (RBCs)
• Liquefaction
• Normal:15–30 minutes after collection
• Lumpy >60 min – sperms may be trapped in unliquefied jelly; maybe
sign of prostatic infection, lack of prostatic protease
• Viscosity
• Normal;Smooth and watery
• Abnormal –, thick with long threads (21G needle). High viscosity
impede sperm movements
Hassaan Ali 20154/7/2015
20. sperm count
• Certain conditions may be linked with a low or absent
sperm count. These conditions include :
• Orchitis
• Varicocele
• Klinefelter syndrome
• Radiation treatment to the testicles
• Diseases that can cause shrinking (atrophy) of the testicles
(such as mumps).
• Long term illness such as diabetes which may cause
retrograde ejaculation
• Hormonal imbalance (testosterone, luteinizing hormone
(LH), follicle-stimulating hormone (FSH), or prolactin
• Azoospermia - A small sample (biopsy) of the testicles may
be needed for further evaluation.
Hassaan Ali 20154/7/2015
21. Motility
• Fertilization of an ova is dependent on the ability of the sperm to reach
and unite with it.
• Grade 4 (A): Sperm with progressive motility. These are the strongest and
swim fast in a straight line.
• Grade 3 (B): Sperm with non-linear forward motility. These also move
forward but tend to travel in a curved or crooked motion.
• Grade 2 (C): These have non-progressive motility because they do not
move forward despite they move their tails .
• Grade 1 (D): These are immotile and fail to move at all .
Hassaan Ali 20154/7/2015
22. Morphology
• Refers to the shape and size of the sperm. A normal
sperm has an oval head and a tail 7- 15 times longer
than the head. Abnormal sperm morphology can be a
contributing factor to infertility.
• Causes of abnormal morphology include;
• Congenital testicle abnormalities.
• Fever.
• If any abnormal morphology is seen the semen analysis
should be repeated in four to two weeks to check if the
changes are temporary or permanent.
.
Hassaan Ali 20154/7/2015
24. Vitality
• Distinguishes between samples with live, but
immotile sperm (e.g Kartagener syndrome),
and those with lots of dead sperm (could
result from: sperm senescence;
• exposure to detergents or lubricants; or
spermotoxic antibodies)
Hassaan Ali 20154/7/2015
25. Other cells in semen
• Leukocytes: normally (1-4/HPF), increase number
(leukocytospermia) indicates reproductive tract infection
• Epithelial cells: normally (1-2/HPF)
• Spermatocytes: (Immature germ cells) 1-2/HPF.
• Erythrocytes: (1-2/HPF). Increased number may indicate a
reproductive tract infection or damage to a small capillary
during sample production.
• Note: bacteria and protozoan such as Trichomonas vaginalis
are uncommon in human semen but their presence is
indicative of possible male reproductive tract infection and
should be reported to the referring doctor for further
evaluation.
Hassaan Ali 20154/7/2015
26. Semen biochemistry
• Acid phosphatase: marker for prostatic function
• Citric acid: can indicate prostatic function – low levels may
indicate dysfunction or a prostatic duct obstruction
• Zinc: marker for prostatic function – colorimetric assay
(WHO)
• Fructose: marker for seminal vesicle function, and is a
substrate for sperm metabolism – spectrophotometric
assay (WHO)
• -Glucosidase: secreted exclusively by the epididymis and
so is a marker for epididymal function – spectrophotometric
assay (WHO)
Hassaan Ali 20154/7/2015
27. • A CASA system is an automated, objective and standardized
equipment that allows to assess all the parameters of
sperm in a semen sample.
• CASA are most often used for the assessment of sperm
concentration and motility following the WHO criteria.
• By use of CASA several specific motility parameters in a
more detailed manner such as velocity can be obtained.
Hassaan Ali 20154/7/2015
28. Advantages of CASA
• Advantages
– More objective and reproducible measurement
– Superior documentation of laboratory values
– Superior in measurement of sperm motility
• Disadvantages
– Not reliable if sperm density is <2x106/ml or >50x106/ml, lots of
debris/immotile sperm. Require dilution if >40 x 106/ml in
isotonic buffer to avoid sperm collision
– Need to record 200 sperms for accurate distribution of velocity.
– Parameters not standardized between laboratories –difficult to
interpret results
– No improvement on the manual method in distinguishing
fertilizing capacity of semen
Hassaan Ali 20154/7/2015