SHRI RAM MURTI SMARAK INSTITUTE OF PARAMEDICAL SCIENCES

Course : BMLT
Year : IIIrd year(2019)
Subject : Pathology
Topic : SEMEN ANALYSIS

Lecture by
Dharmendra singh
 Contents
Definition
Formation & Production
Composition
Sample Collection/Patient Preparation
Methods of Collection
Purpose of Semen Analysis
Analytical Methods- Physical, Chemical & Microscopic
Factor Affecting of Semen Analysis
Definition:- Semen is also known as seminal fluid , is an
organic fluid containing spermatozoa. It is formed by the
gonads(testis) as well as accessory male reproductive organ.
Semen is typically translucent with White ,grey ,or even
yellowish tint .
During the process of ejaculation , sperm passes through the
ejaculatory duct and mixed with fluids from seminal
vesicles,the prostate and the bulbourethral glands to form the
semen.

Formation and production--The seminal vesicles produce a

yellowish viscous fluid that is rich in fructose and other
substance that makes up about 70% human semen.
Approximately 70% of the semen volume is derived from seminal
vesicles this fluid is neutral or slightly alkaline and appeared as pale
yellow colour due to containing the flavin.

Semen is a composite solution basically consists of spermatozoa

suspended in the seminal plasma. Spermatozoa (sperm)are the male
sex cells present in semen that carry a man's genetic material and
constitute less than 5% of the semen volume .

spermatozoa are largely stored in the ampullary portions of

vas deferens until released in the process of ejaculation. it is
formed by the testis.
About 20% of the volume of plasma is contributed by prostate gland .this
fluid is slightly acidic pH 6.5 due to high concentration of citric acid. it is
milky appearance and also rich in proteolytic enzymes and
acid phosphatase.

These proteolytic enzymes responsible for coagulation and liquefaction

of semen.

About 10 to 15% of the semen volume is also contributed by epididymis,

vas deferens and urethral glands. Epididymis secrete a number proteins
into the tubular lumen that are essential for fertilizing capacity of
sperms.
COMPOSITION OF SEMEN
In 1992 WHO describe normal semen having physical and
chemical properties.
1.--volume........... 2-6 ml
2--PH...................7.2-8.0
3--viscosity........... Fairly viscous when ejaculated but liquefy in
30 minutes.
4--motility............within 2 hours of ejaculation 60 to 80% of
sperm are vigorously motile. In 6 to 8 hours 22 to 35% still
motile
5-osmolarity......... 354 m. osmol.
 Property Per 100mL In average volume (3.4 mL)
Calcium (mg). 27.6 0.938
Chloride (mg) 142 4.83
Citrate (mg) 528 18.0
Fructose (mg). 272 9.25
Glucose (mg) 102 3.47
Lactic acid (mg) 62 2.11
Magnesium (mg) 11 0.374
Potassium (mg) 109 3.71
Protein (g) 5.04 0.171
Sodium (mg) 300 10.2
Urea (mg) 45 1.53
Zinc (mg) 16.5 0.561
Sample collection .........Patient preparation
Ask the patient may be avoid any sexual activity in 2 to 5 days
before analysis that help to ensure the result
If the patient avoid sexual activity more than 2 weeks before the
test because a long gapping between sexual activity to form more
and more active sperm.
Ask the patient to avoid drinking alcohol or any type of medicine
Methods of collection
It is very important to relax the patient and allow the production
of specimen. To take place in surrounding which are most
comfortable emotionally and physically.
The patient should be allowed if necessary to select where he wishes
to produce the specimen .
 It may be at home or in a private room of laboratory or hospital
which is for away from other people, roads etc; very quiet which
will guarantee total privacy.
The optimal sexual abstinence for semen sample obtaining is from
5 to 7 days.
 Different methods used for semen collection are masturba-
tion, condom collection and epididymal extraction, etc.
The most common way of collection of semen by masturbation
that is complete and uncontaminated. Specimen may be
collected by coitus interruptus for those patient who are
reluctant to produce semen by masturbation.
Semen produced by masturbation in which during sexual
intercourse the penis from his partner just before ejaculation.
The patient can also collect the semen sample during sex by
using condom (male factor pack)
Specimen collect in clean dry wide mouth plastic containers
(50ml).
Specimen transport to the laboratory within 2 hours of
ejaculation . keep away from exposure to direct sunlight or
excessive cold , if collection proceed at home, office, clinic
that is close to hospital and transport easily.
SEMEN ANALYSIS
Following are the various important purposes off routine
semen analysis:
evaluation of infertility
Artificial insemination
to checkout the vasectomy
In medico legal cases example sexual claim of violence rape
examination of stored semen specimen (may be in the case of a
husband away from home for a long period and when the wife is
undergoing complicated infertility therapy ).
for men whose future fertility is threatened may be.
Methods for examination of semen
PHYSICAL EXAMINATION.. including
Colour, volume, appearance, liquifacation time, viscosity, Ph ,
odour
1.Colour-semen is normally whitish ,greay- white or slightly
yellowish.
Presence of blood semen become pink or red( Hematospermia)
Presence of bilirubin may impart yellow color
Presence of urine high urea content that confirm presence of
urine
2. Volume-2-6 ml semen ejaculate
Alteration
Low volume-due to abnormalities in vinyl cycle or prostate
gland and various neurological disorder.
Some dietary supplements have been marketed with claims to
increase seminal volume. Like other supplements, including
so-called herbal viagra.
Aspermia- total absence of ejaculate
Oligo or hypospermia- reduction in the volume of
ejaculate
Hyperspermia- increase in semen volume
3. Viscosity-when ejaculated , semen is fairly viscous .
To check out viscosity of semen falls drop by drop that indicate
the normal viscosity of semen
Increase viscosity affect motility of sperm

4. Liquefaction time-15 to 30 minute(liquefaction occurs

because of presence of fibrinolysin)
If increase due to infection
Due to defect of proteolytic enzyme
Note :The liquefaction is the process when the gel formed by
proteins from the seminal vesicles is broken up and the
semen becomes more liquid. It normally takes less than 20
minutes for the sample to change from a thick gel into a
liquid. In the NICE guidelines, a liquefaction time within 60
minutes is regarded as within normal ranges.

5. Odour-musti odour

Semen normally smells like ammonia, bleach, or chlorine.

6. Appearance-Semen is typically translucent with white,
grey or even yellowish tint. Blood in the semen can cause a
pink or reddish colour, known as hematospermia.

7. PH-7.2 to 8.0

Note:Acidic ejaculate (lower pH value) may indicate one or

both of the seminal vesicles are blocked. A basic ejaculate
(higher pH value) may indicate an infection. A pH value
outside of the normal range is harmful to sperm and affect
their ability to penetrate the egg.
FACTOR AFFECTING OF SEMEN ANALYSIS
Alcohol, heroin, smoking, tobacco , that influence the result
Herbal medicine like as Viagra that influence the volume of semen.
Coldness that increase the motility of sperm
NOTE : Compared to samples obtained from masturbation, semen
samples from collection condoms have higher total sperm counts,
sperm motility, and percentage of sperm with normal
morphology[citation needed]. For this reason, they are believed to
give more accurate results when used for semen analysis.
If the results from a man's first sample are subfertile, they
must be verified with at least two more analyses. Atleast 2
to 4 weeks must be allowed between each analysis

CHEMICAL EXAMINATION
Fructose concentration test
determine the amount of energy available to the semen for
moving. if fructose concentration is decrease then the motility
and total sperm concentration will be decrease.
Fructose is measured qualitatively by -resorcinol test -------
Procedure--5 ml resorcinol reagent+0.5 ml semen+heat to
boil and observe the colour
Red colour indicate presence of fructose
No colour indicate deficiency of fructose that is occurred due
to in androgen deficiency or ejaculatory obstruction of semen
Microscopic examination -
Motility- wet mount preparation after liquefied semen
Procedure--place a drop of liquefied semen on slide, and
cover it with cover slip ,the entire drop of semen and examine
under microscope
Count the number of actively motile sperms out the total counts of
200...
 Vigorously motile sperm --60 to 80%
 Sluggish motile sperm-- 20 to 30%
 Non motile sperm--------- 0 to 10%

2. Total sperm count --- normal sperm count 20 to 60 million/ ml
This is done in Neubauer's chamber using a WBC pipette .
semen diluting fluid
Composition----sodium bicarbonate-----------------5 gm
Formalin --------------------------------- 1 ml
Distilled water--------------------------99 ml
Procedure--draw liquified semen in wbc pipette upto 0.5
mark and then draw diluting fluid upto 11 mark
 Mix well, load chamber and allowed the spermatozoa to settle
for 2 minut
Examine under microscope and count the spermatozoa as WBCs
Calculation:-
sperms/ml of semen = N×50×1000.
( N mean No.of Sperm )
3. Determination of morphology of sperm
Staining procedure
Prepare a thin smear from liquefied semen on a glass slide .
Let it dry in the air and then heat very gently to fix .
Stain the smear by using leishman's stain.
Rinse the slide and dry.
Examine under the microscope.
Observe at least 200 spermatozoa for any abnormalities in the
morphology .
Normally 60- 80% of spermatozoa are normal and about 20
% of spermatozoa are in abnormal forms ...
These are following as :
Normal sperm have an oval head with a long tail.
 Abnormal sperm have head or tail defects — such as a large
or misshapen head or a crooked or double tail.
These defects might affect the ability of the sperm to reach
and penetrate an egg.
 TERMS
Aspermia:absence of semen.
Azoospermia:describe a total absence if spermatozoa in semen.(After cen-
trifuge sperm count is zero/hpf).
Oligozoospermia:refers to a reduced no. of spermatozoa in semen and is us-
ally used to describe a sperm conc. Of less than 20 million/ml.Sperm count 5-
10 sperm/HPF.
Severe oligospermia:Sperm count 1-2 sperm/hpf.
Polyzoospermia:Denotes an increase no.of spermatozoa in semen and is usu-
ally refers a sperm conc.in excess of 350million/ml.
THANK U……………….