Evaluation of semen
1-Volume
The graduated collection tube was utilized to determine the volume of
ejaculate to the nearest 0.1ml, just after collection.
2-Hydrogen ion concentration (pH) and color:
Values of seminal pH determined by using pH paper.
3-Motility
a)Mass motility
A drop of fresh semen was deposited on a warm glass slide (37°C) and its
motility was evaluated according to wave motion table under microscope .
Table : Evaluation of mass motility.
%
Wave motion
0
Total immobility
10
Individual movement
20-40
Very slow movement
45-65
General wave motion ,slow amplitude of waves
70-85
Rapid wave motion ,no eddies
90-100
Rapid wave motion, eddies
b) Individual motility
Individual sperm motility was evaluated by depositing a few drops of diluted
semen by normal saline on a warm glass slide (37°C), covered by cover
slip(22mmX22mm). Evaluation was made according to progressively foreword
motility as in Table under microscope .
Table : Evaluation of sperm individual motility
%
Individual motility
0
Total immobility
10
1/5 of sperms motile
20-40
2/5 of sperms motile
45-65
3/5 of sperms motile
70-85
4/5 of sperms motile
90-100
All sperms motile
4-Dead sperms percentage:
A drop of fresh semen was mixed with one drop of eosin stain then with two
drops of nigrosin stain and smear made and dried. The slide was examined under
microscope, uncolored sperm head(white) is considered as life sperm ,while pink
head(colored) due to dead sperm .
5-Abnormal sperms percentage:
Determination of abnormal sperm percentage was made using the same
smear of live and dead.
6-Sperm concentration:
Estimation of sperm concentration was made by adding 0.1ml of fresh
semen to 19.9 ml of sperm counting solution (2.9 sodium citrate +few drops of
eosin to facilitate distinguishing of sperm under microscope +Mercuric chloride
(Hgcl2) as killing agent for sperm). The solution components were mixed for 3
minutes, after that using Neubauer chamber (RBCs square) sperms were counted
to determine sperm concentration using the following equation:
C=n X200 X 400 X 10 X1000/ 80
-n=No .of sperm in five square.
-200= converse dilution rate.
-80=No .of small square in five large squares.
-10= converse depth between slide and cover slide.
-400= converse square area of cover slide.
Sperm function tests
Semen analysis provides enough information to
recognise sires of very low fertility, but has been
increasingly considered to be a poor discriminator
between moderate and high fertility levels
. In order to attempt to improve
the accuracy of semen assessments, a number of
tests of sperm function have been employed, with
varying success. The simplest of such tests incubate
semen at various temperatures (typically 4 or
40°C) and, by relating the duration of sperm
survival under these conditions to survival in the
female genital tract, produce reasonable correlations
with fertility. Other tests
utilise additional measurements upon the semen,
such as pH, adenosine triphosphate (ATP) content
or aspartate transaminase concentration
. These have been moderately
successful, but have not been of sufficiently
greater value than conventional semen assessment
to justify their use. In medical practice, much
value is placed upon the ability of sperm to penetrate
cervical mucus and the behaviour of the
sperm at the interface between semen and mucus
. Failure of mucous
penetration is frequently a sign of failure of sperm
function and occurs in sperm that have been
damaged by cryopreservation or in the presence
of anti-sperm antibodies.
Of more widespread use in veterinary practice is
computer-assisted analysis of sperm swimming
characteristics. In medical practice, such analyses
are regarded as a useful prognostic tool in assessment
of fertility because high correlations have
been demonstrated between such measurements
and fertility. The most important swimming characteristics
are rate of forward progress, lateral
movement of the sperm head and characteristics
of the flagellar beat. Although the use of sperm
motility analysis in veterinary practice at present is
largely confined to thoroughbred stallions
and AI stud bulls, it
is probable that the use of such systems will rapidly
increase as the cost of analysis programs decreases.
Assessments of sperm viability have also been
improved in recent years. Fluorescent markers
that stain live, but not dead, sperm have been used
and high correlations with fertility demonstrated
. Assessment of the proportion
of sperm with intact acrosomes has been
highly correlated with fertility .
The most recent innovation in assessment of
sperm function has derived from the development
of in vitro fertilisation (IVF) procedures,
in which sperm from different sires were observed
to have widely differing fertilisation success rates.
Subsequently, the ability of sperm to undergo
acrosome reaction in vitro was identified as a critical
stage in the IVF procedure and, in the bull,
tests of sperm function based upon in vitro induction
of acrosome reactions have been found to
have very high correlation with fertility in the field