Activity 9 Semenalysis

ST.
ALEXIUS COLLEGE
Gen. San. Drive, City of Koronadal, South Cotabato, Philippines 09506, Tel.: (083) 228-2019, Fax: (083) 228-4015, Email: st.alexiuscollege@yahoo.com
ACTIVITY 9
SEMEN ANALYSIS
I. INTRODUCTION
A high quality, comprehensive semen assessment is not just the cornerstone of the
diagnosis of male subfertility, it is also the starting point for providing prognostic information.
Male subfertility poses a significant global challenge, with epidemiological evidence indicating
that approximately one in seven couples experience subfertility. Among the various causes,
sperm dysfunction stands out as the most prevalent contributor to male subfertility.
Recent advancements in andrology and assisted reproductive technology (ART), coupled

with the growing awareness of fertility issues, particularly among couples choosing to conceive
later in life, have led to an increased focus on semen analysis.
Semen analysis offers a comprehensive evaluation of the male partner's reproductive

function within a subfertile couple. This assessment encompasses various aspects, including
sperm counts, which reflect sperm production, transport through the male reproductive system,
and ejaculatory function. Additionally, assessments of sperm motility serve as a fundamental
indicator of sperm viability and competence. The evaluation also considers sperm vitality,
distinguishing between dead and live but immotile spermatozoa, as well as sperm form, which
provides insights into aspects of sperm production and maturation. The physical characteristics
of the ejaculate, such as semen volume, are also examined during semen analysis.
A thorough and high-quality semen assessment not only serves as the cornerstone for
diagnosing male subfertility but also provides valuable prognostic information for the couple's
fertility journey. It offers essential insights into the male partner's reproductive health, assisting
in determining the appropriate treatment approach and predicting the likelihood of successful
conception.
II. OBJECTIVES
At the end of the activity, students must be able to:

1. perform the different steps in semen analysis with accuracy and precision,
2. interpret the result of the semen analysis,
3. report properly the result of the semen analysis, and
4. answer correctly the research questions.
Clinical Microscopy Laboratory Manual 91

Compiled by: Carl Jethro G. Dellosa, RMT & Janine Teza S. Villena, RMT, MSMT
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III. MATERIALS
Screw-capped sterile transparent plastic container

Record sheet
Automatic pipette
Pipette tips
Dropper
Plain test tubes
Test tube racks
Timer
pH test strip
Glass slides
Coverslips (22 x 22 mm)
Microscope
Immersion oil
Cell counter
Differential cell counter
Marker
Chilled distilled water
Neubauer counting chamber with cover slips
RBC Thoma pipette
Improvised suction device for Thoma pipette
Methanol
Wright’s stain (Eosin and Methylene Blue)
Slide rack
Forceps
Calculator
Disinfectant
Centrifuge
Tissue
IV. PROCEDURE
PREANALYTICAL PHASE
A. Patient Instructions for Sample Collection, Safe Handling and Delivery

1. Patient should be given clear and simple instructions explaining the need for semen
analysis and what is required for specimen collection.
2. Patient should be informed about the importance of abstinence time. Ejaculate must be
collected after 3-5 days (but not more than 7 days) of abstinence.

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3. Samples should be obtained by masturbation and collected in a warm (20-40ºC), sterile,

nontoxic plastic or glass wide-mouth container. Prior to sample collection, the patient
must void and wash hands and genitals to minimize the chances of contamination.
4. Use of lubricants and saliva should be avoided as their potential toxicity might influence
the result. Semen samples should be protected from extremes of temperature (<20ºC or
>40ºC) during transport to the laboratory.
5. All sample containers are labeled with adequate information to eliminate any chances of
error.
6. Regular condoms should not be used because of their spermicidal effect. Ideally, the
samples must be collected close to the laboratory. If the specimen cannot be produced
close to the laboratory, it must be delivered to the laboratory as soon as possible, certainly
within 1 hour of collection. During this period, the sample has to be kept warm by
carrying it next to the body, and temperature extremes must be avoided.
B. Information on the Sample Container and Patient Sheet

Once the patient has collected the specimen, some preliminary information about the
specimen should be obtained:
1. Label the specimen clearly, indicating patient’s complete name, clinic number, and
collection date.
2. Record collection date and time.
3. Record abstinence time in days.
4. Record time the sample is received at the laboratory.
5. Record information about split ejaculate, noting when the sample was lost and
whether the spill occurred at the beginning or after ejaculation. The samples always
should be labeled as “Biohazard” and extreme precaution should be taken. Follow
safety guidelines and protocols in handling the sample as the semen sample may
contain infectious agents (e.g. hepatitis B, hepatitis C, HIV, herpes simplex).
ANALYTICAL PHASE
A. Semen Volume and Handling of the Specimen Container

1. Weigh the container after the ejaculate has been collected.
2. Calculate the specimen volume by subtracting the tare weight from the result of weighing
after specimen collection.
3. Record the value on the form. Time of ejaculation is written clearly on the vial.
4. Place the specimen container immediately in the incubator and record the time on the form.
Leave in the incubator until 25-30 minutes after ejaculation.
Reference:

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Normal Volume: 2-5 mL
B. Semen Appearance and Odor

1. Open the lid of the container.
2. Evaluate the color and smell of the semen.
3. Record the observation on the form.
Reference:
Normal color: gray-white, translucent
Odor: musty
C. Semen Liquefaction
1. Record the liquefaction time of semen. Duration begins from the time of collection.
If after 2 hours the specimen has not liquefied, proteolytic enzymes such as alpha-
chymotrypsin may be added to allow the rest of the analysis to be performed.
Reference:
Normal: 30-60 minutes
D. Semen Viscosity
1. Aspirate the semen into a 5 mL plastic volumetric pipette.
2. Let the semen run out of the pipette back into the sample collection container.
3. Estimate the length of the threads formed between the droplets.
4. Record the result.
Reference:
Normal: threads formed 20mm and below or pours in droplets.
If threads formed are >20 mm long when the semen drops from the pipette, the sample
is considered to have an abnormal viscosity. This finding should be recorded on the
report form. The linger the thread the more sever the abnormal viscosity.
E. Semen pH
1. Transfer a small droplet of semen onto the test strip using a pipette.
2. Allow the color to develop.
3. Compare the color of the test strip with the appropriate color scale to read the pH.
4. Record the result on the form.
Reference:
Normal pH: 7.2-8.0

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F. Sperm Motility
I. Making a Wet Preparation
1. Use a clean microscope slide that has been pre-warmed to 370C. Store the slide warm
during the making of the preparation and ideally keep it at 370C during the entire
investigation of the preparation.
2. Swirl the semen before taking the aliquot for the wet preparation to make sure the
specimen is well mixed.
3. Aspirate the appropriate volume using a pipette and deposit it as a droplet in the
middle of the microscope slide.
4. Apply the coverslip (22 x 22 mm coverslip) to the droplet immediately.
II. Initial Evaluation of a Wet Preparation

Note: The examination of the wet preparation should begin as soon as the flow has
ceased, which should happen within 1 minute after the coverslip on the semen droplet; if
not, then the preparation should be discarded and a new one should be prepared.
1. Count the number of spermatozoa visible in randomly chosen microscope fields. If
less than one spermatozoon is observed per “high power field”, the sample should be
treated as an azoospermic or severely oligozoospermic sample.
2. Assess the spermatozoa motility if spermatozoa are observed.
3. Assess the presence of sperm aggregation and agglutination: The degree of sperm
aggregation or agglutination is determined in 10 randomly chosen microscope fields
not adjacent to the coverslip edges. The proportion of spermatozoa trapped in clumps
is estimated to the nearest 5%.
• Agglutination is defined as at least some motile spermatozoa adhering without
other cells and debris, most likely due to the presence of multivalent anti-sperm
antibodies. If the agglutinates are very large, it may be difficult to assess whether
the spermatozoa are specifically (e.g. head-to-head or tail-to-tail) or more
randomly bound to each other.
• Clumps comprising immotile (dead) spermatozoa and debris are described as
aggregation. Small aggregates are common in normal samples, while large
aggregates, which can contain hundreds of spermatozoa each, are considered as
abnormal.
• In samples where the proportion of spermatozoa involved in clumping is
estimated to be 25% or more, motility is assessed on the free spermatozoa only.
The proportion of spermatozoa involved in clumping, and a note that motility was
assessed on the free spermatozoa only, must be recorded on the specimen form.
4. Assess the presence of other cells and debris.
It is common to find other cells and debris in semen samples. Therefore, only the
increased presence of such material should be reported if seen in multiple fields. Such

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abnormalities should be commented upon in the report form using a few different,
standardized expressions:
• Round cells: These are common in semen specimens. “Leukocytes” must be
differentiated from immature germinal epithelium cells or precursors, or large cell
bodies (often without nuclei) consisting of cytoplasm that has become exfoliated
from the seminiferous epithelium of the testis. Other round cells present in semen.
Round cells can be counted in the same fields used for motility assessment or,
alternatively, in Neubauer hemocytometer chamber at the same time as the
assessment of sperm concentration.
III. Motility Testing

1. Count the rapid and slow progressive spermatozoa, and then count the non-
progressive and immotile spermatozoa in the same microscope field. If the
concentration of spermatozoa is very high, it is advisable to only count spermatozoa
in a smaller field, e.g. in the area of four central squares in an eyepiece grid.
2. Assess at least five different fields, and count at least 200 spermatozoa in each
preparation.
3. Repeat the assessment of the motility of at least 200 spermatozoa in a second wet
preparation from the same semen specimen.
4. Calculate:
a. Proportions for the four categories (rapid, slow progressive, non-progressive
and immotile) for each of the two preparations
Formula:
𝑃𝑟𝑜𝑝𝑜𝑟𝑡𝑖𝑜𝑛 𝑓𝑜𝑟 𝑒𝑎𝑐ℎ 𝑐𝑎𝑡𝑒𝑔𝑜𝑟𝑦
𝑇𝑜𝑡𝑎𝑙 𝑁𝑜. 𝑜𝑓 𝑠𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑖𝑛 𝑒𝑎𝑐ℎ 𝑚𝑜𝑡𝑖𝑙𝑖𝑡𝑦 𝑔𝑟𝑜𝑢𝑝
=
𝑇𝑜𝑡𝑎𝑙 𝑁𝑜. 𝑜𝑓 𝑠𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑎𝑠𝑠𝑒𝑠𝑠𝑒𝑑 𝑖𝑛 𝑡ℎ𝑎𝑡 𝑝𝑟𝑒𝑝𝑎𝑟𝑎𝑡𝑖𝑜n

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b. Average proportion of duplicate slides per category
Formula:
𝑆𝑙𝑖𝑑𝑒 1 + 𝑆𝑙𝑖𝑑𝑒 2
𝐴𝑣𝑒𝑟𝑎𝑔𝑒 𝑝𝑟𝑜𝑝𝑜𝑟𝑡𝑖𝑜𝑛 𝑜𝑓 𝑑𝑢𝑝𝑙𝑖𝑐𝑎𝑡𝑒𝑠 𝑝𝑒𝑟 𝑐𝑎𝑡𝑒𝑔𝑜𝑟𝑦 =
2
c. Difference between duplicate slides per category
Formula:
𝐷𝑖𝑓𝑓𝑒𝑟𝑒𝑛𝑐𝑒 𝑏𝑒𝑡𝑤𝑒𝑒𝑛 𝑑𝑢𝑝𝑙𝑖𝑐𝑎𝑡𝑒 𝑠𝑙𝑖𝑑𝑒𝑠
= 𝐷𝑜𝑚𝑖𝑛𝑎𝑛𝑡 𝑆𝑙𝑖𝑑𝑒 − 𝑁𝑜𝑛𝑑𝑜𝑚𝑖𝑛𝑎𝑛𝑡 𝑆𝑙𝑖𝑑𝑒
Note: All motility results are presented as integer percentage values. The sum of the four
motility categories (a,b,c,d) in a sample should be 100. If, due to rounding errors, the sum is
99 or 101, adjust the dominant group to get a sum of 100.
Reference:
Normal: >50% within 1 hour
G. Sperm Concentration and Sperm Count

1. Prepare a clean plain tube.
2. Aspirate 950 uL of diluent (chilled distilled water) into the plain tube.
3. Aspirate 50 uL of a well-mixed, liquefied ejaculate into the plain tube.
4. Tap gently the mixture (1:20 dilution).
5. Load in both hemocytometer counting chambers.
6. Allow to settle for 2 minutes.
7. Count the sperm in the four corner center squares of the large center square. The counts should
agree within 10%. If the counts do not agree, both the dilution and the counts are repeated.

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• Only fully developed sperm should be counted.

• Immature sperm and WBCs, often referred to as “round” cells, must not be included.
• >1,000,000/mL WBCs is associated with inflammation or infection of the reproductive
organs that can lead to infertility.
• >1,000,000/mL spermatids indicate disruption of spermatogenesis. This may be caused by
viral infections, exposure to toxic chemicals, and genetic disorders.

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8. Calculate the sperm concentration.

Formula:
𝑆𝑝𝑒𝑟𝑚 𝐶𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛 (𝑚𝑖𝑙𝑙𝑖𝑜𝑛 𝑝𝑒𝑟 𝑚𝐿)
= 𝑆𝑝𝑒𝑟𝑚 𝐶𝑜𝑢𝑛𝑡𝑒𝑑 (𝑎𝑣𝑒𝑟𝑎𝑔𝑒 𝑜𝑓 2 𝑐ℎ𝑎𝑚𝑏𝑒𝑟𝑠)𝑥 1,000,000
Reference:
Normal: >20 million/mL
9. Calculate the sperm count.
Formula:
𝑆𝑝𝑒𝑟𝑚 𝐶𝑜𝑢𝑛𝑡 (𝑚𝑖𝑙𝑙𝑖𝑜𝑛 𝑝𝑒𝑟 𝑒𝑗𝑎𝑐𝑢𝑙𝑎𝑡𝑒)
= 𝑆𝑝𝑒𝑟𝑚 𝐶𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛 (𝑝𝑒𝑟 𝑚𝐿) 𝑥 𝑆𝑝𝑒𝑐𝑖𝑚𝑒𝑛 𝑉𝑜𝑙𝑢𝑚𝑒
Reference:
Normal: >40 million/ejaculate
H. Sperm Morphology
1. Prepare two clean glass slides.
2. Put 10uL of sperm from a well-mixed specimen. If the sperm concentration is very low (<2 x
106/mL) an aliquot of the semen specimen can be centrifuged (maximum 1000 g) and the pellet
resuspended in a small amount of clean seminal plasma (i.e. the supernatant) to make the
morphology smear.
3. Prepare a smear.
Diagram illustrating two alternative techniques for making smears for sperm morphology
assessment.

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4. Air dry slides.

5. Fix slides in methanol.
6. Air dry slides.
7. Dip in Eosin stain for 5 to 6 seconds.
8. Wash slides in running water.
9. Dip in Methylene Blue stain for 10-30 seconds.
10. Wash with running water.
11. Air dry slides.
12. Load the stained smear on the microscope and scan the smear with a low power objective to
observe the spreading of the spermatozoa, the staining quality, and the presence of round cells.
If round cells are observed, move to 40x objective to identify the type of round cells present.
13. Identify suitable area for the performance of the sperm morphology evaluation.
14. Add a small drop of immersion oil on the slide where the light is shining through it, without
removing the slide from the microscope stage.
15. Bring the oil immersion objective into place.
16. Examine the smear under oil-immersion objective.
17. Assess at least 200 spermatozoa. Record the morphological appearance of each spermatozoon as
“ideal” or “abnormal”. Assign four buttons on the counter for each of the four types of defects
(head, neck/midpiece, tail, presence of cytoplasmic residues).
Normal Spermatozoa Structure

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Abnormalities of Sperm Heads and Tails
18. Compute for the value of normal and abnormal forms.

Formula:
𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝐹𝑜𝑟𝑚 (%)
= 𝑁𝑜. 𝑜𝑓 𝑛𝑜𝑟𝑚𝑎𝑙 𝑠𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑐𝑜𝑢𝑛𝑡𝑒𝑑 𝑥 𝑆𝑒𝑝𝑐𝑖𝑚𝑒𝑛 𝑉𝑜𝑙𝑢𝑚𝑒
𝑇𝑜𝑡𝑎𝑙 𝑁𝑜.𝑜𝑓 𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝑆𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑐𝑜𝑢𝑛𝑡𝑒𝑑
𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝐹𝑜𝑟𝑚 (%) = 200
x 100
𝑇𝑜𝑡𝑎𝑙 𝑁𝑜. 𝑜𝑓 𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝑆𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑐𝑜𝑢𝑛𝑡𝑒𝑑

𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝐹𝑜𝑟𝑚 (%) = x 100
200
Reference:
Normal (Routine Criteria): >30% normal forms
Normal (Strict Criteria): >14% normal forms

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I. Calculation of Round Cells

1. Count the number of spermatids or leukocytes seen in conjunction with 100 mature sperm.
2. Calculate the amount of round cells.
Formula:
𝑁x𝑆
𝐶=
100
C = round cell concentration

N = number of spermatids or neutrophils counted per mature sperm
S = sperm concentration in million per milliliter
Reference:
Normal: <1.0 million/mL
POST ANALYTICAL PHASE

1. Record the results on the result form.
2. Dispose wastes into appropriate bins.
3. Remove PPE and wash hands.

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ACTIVITY 9
SEMEN ANALYSIS
Performance Assessment Sheet
Name:___________________________________________ Date:______________________
PROCEDURE RATING COMMENTS

1. Wash hands and wear PPE.
PREANALYTICAL PHASE
1. Orient patients thoroughly of the instructions,
sample collection, safe handling, and deliver of
specimen.
2. Label the specimen clearly, indicating patient’s
complete name, clinic number, and collection date.
3. Record collection date and time.
4. Record abstinence time in days.
5. Record time the sample is received at the
laboratory.
6. Record information about split ejaculate, noting
when the sample was lost and whether the spill
occurred at the beginning or after ejaculation. The
samples always should be labeled as “Biohazard”
and extreme precaution should be taken. Follow
safety guidelines and protocols in handling the
sample as the semen sample may contain
infectious agents (e.g. hepatitis B, hepatitis C,
HIV, herpes simplex).
ANALYTICAL PHASE
A. Semen Volume and Handling of the Specimen
Container
1. Weigh the container after the ejaculate has been
collected.
2. Calculate the specimen volume by subtracting the
tare weight from the result of weighing after
specimen collection.
3. Record the value on the form. Time of ejaculation
is written clearly on the vial.
4. Place the specimen container immediately in the
incubator and record the time on the form. Leave
in the incubator until 25-30 minutes after
ejaculation.

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B. Semen Appearance and Odor

1. Open the lid of the container.
2. Evaluate the color and smell of the semen.
3. Record the observation on the form.
C. Semen Liquefaction
1. Record the liquefaction time of semen. Duration
begins from the time of collection. If after 2 hours
the specimen has not liquefied, proteolytic
enzymes such as alpha-chymotrypsin may be
added to allow the rest of the analysis to be
performed.
D. Semen Viscosity
1. Aspirate the semen into a 5 mL plastic volumetric
pipette.
2. Let the semen run out of the pipette back into the
sample collection container.
3. Estimate the length of the threads formed between
the droplets.
E. Semen pH
1. Transfer a small droplet of semen onto the test strip
using a pipette.
2. Allow the color to develop.
3. Compare the color of the test strip with the
appropriate color scale to read the pH.
4. Record the result on the form.
F. Sperm Motility
I. Making a Wet Preparation
1. Use a clean microscope slide that has been
pre-warmed to 370C. Store the slide war,
during the making of the preparation and
ideally keep it at 370C during the entire
investigation of the preparation.
2. Swirl the semen before taking the aliquot
for the wet preparation to make sure the
specimen is well mixed.
3. Aspirate the appropriate volume using a
pipette and deposit it as a droplet in the
middle of the microscope slide.
4. Apply the coverslip (22 x 22 mm coverslip)
to the droplet immediately.
II. Initial Evaluation of a Wet Preparation

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5. Count the number of spermatozoa visible in

randomly chosen microscope fields. If less
than one spermatozoon is observed per
“high power field”, the sample should be
treated as an azoospermic or severely
oligozoospermic sample.
6. Assess the spermatozoa motility if
spermatozoa are observed.
7. Assess the presence of sperm aggregation
and agglutination: The degree of sperm
aggregation or agglutination is determined
in 10 randomly chosen microscope fields
not adjacent to the coverslip edges. The
proportion of spermatozoa trapped in
clumps is estimated to the nearest 5%.
8. Assess the presence of other cells and
debris.
III. Motility Testing
1. Count the rapid and slow progressive
spermatozoa, and then count the non-
progressive and immotile spermatozoa in
the same microscope field. If the
concentration of spermatozoa is very high,
it is advisable to only count spermatozoa in
a smaller field, e.g. in the area of four
central squares in an eyepiece grid.
2. Assess at least five different fields, and
count at least 200 spermatozoa in each
preparation.
3. Repeat the assessment of the motility of at
least 200 spermatozoa in a second wet
preparation from the same semen
specimen.
4. Calculate the following:
a. Proportions for the four categories
(rapid, slow progressive, non-
progressive and immotile) for each
of the two preparations
b. Average proportion of duplicate
slides per category
c. Difference between duplicate
slides per category
G. Sperm Concentration and Sperm Count

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10. Prepare a clean plain tube.

11. Aspirate 950 uL of diluent (chilled distilled water)
into the plain tube.
12. Aspirate 50 uL of a well-mixed, liquefied ejaculate
into the plain tube.
13. Tap gently the mixture (1:20 dilution).
14. Load in both hemocytometer counting chambers.
15. Allow to settle for 2 minutes.
16. Count the sperm in the four corner center squares
of the large center square. The counts should agree
within 10%. If the counts do not agree, both the
dilution and the counts are repeated.
17. Calculate the sperm count.
H. Sperm Morphology
1. Prepare two clean glass slides.
2. Put 10uL of sperm from a well-mixed specimen. If
the sperm concentration is very low (<2 x 106/mL)
an aliquot of the semen specimen can be
centrifuged (maximum 1000 g) and the pellet
resuspended in a small amount of clean seminal
plasma (i.e. the supernatant) to make the
morphology smear.
3. Prepare a smear.
4. Air dry slides.
5. Fix slides in methanol.
6. Air dry slides.
7. Dip in Eosin stain for 5 to 6 seconds.
8. Wash slides in running water.
9. Dip in Methylene Blue stain for 10-30 seconds.
10. Wash with running water.
11. Air dry slides.
12. Load the stained smear on the microscope and scan
the smear with a low power objective to observe
the spreading of the spermatozoa, the staining
quality, and the presence of round cells. If round
cells are observed, move to 40x objective to
identify the type of round cells present.
13. Identify suitable area for the performance of the
sperm morphology evaluation.

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14. Add a small drop of immersion oil on the slide

where the light is shining through it, without
removing the slide from the microscope stage.
15. Bring the oil immersion objective into place.
16. Examine the smear under oil-immersion objective.
17. Assess at least 200 spermatozoa. Record the
morphological appearance of each spermatozoon
as “ideal” or “abnormal”. Assign four buttons on
the counter for each of the four types of defects
(head, neck/midpiece, tail, presence of cytoplasmic
residues).
18. Compute for the value of normal and abnormal
forms.
I. Calculation of Round Cells
1. Count the number of spermatids or leukocytes seen
in conjunction with 100 mature sperm.
2. Calculate the amount of round cells.
POST ANALYTICAL PHASE
1. Record the results on the result form.
2. Dispose wastes into appropriate bins.
3. Remove PPE and wash hands.
Rating: 4 = Excellent 3 = Very Satisfactory 2 = Satisfactory 1 = Poor 0 = Not

Done
Level of Competency:
Instructor: Date Evaluated:
Student:

ST. ALEXIUS COLLEGE
ACTIVITY 9
SEMEN ANALYSIS
Report Sheet
Name: _______________________________________ Date: _________________

Score: ________________
RESULT FORM
Complete Name:
Age/Sex:
Date of Birth:
Date of Examination:
Time of Collection:
Time of Submission:
SEMEN ANALYSIS
Parameters Result Normal Value
Volume: 2-5 mL
Appearance: gray-white, translucent
Odor: musty
threads formed 20mm and below or
Viscosity:
pours in droplets
Liquefaction Time: 30-60 minutes
pH: 7.2-8.0
Motility: >50% within 1 hour
Concentration: >20 million/mL
Count: >40 million/ejaculate
Routine Criteria: >30% normal forms
Morphology:
Strict Criteria: >14% normal forms
Round cells: <1.0 million/mL
Performed by:
Validated by:

ST. ALEXIUS COLLEGE
1. Label and give the functions of each part of the sperm cell.
Part Function(s)
1.
2.
3.
4.
5.

ST. ALEXIUS COLLEGE
6.
7.
8.
2. Printed pictures of the step-by-step process of semen analysis.

(Use separate bond papers and attach after this page.)

ST. ALEXIUS COLLEGE
RESEARCH QUESTIONS:
1. Give two latest advancements in sperm analysis and discuss each briefly.
_______________________________________________________________________________
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_______________________________________________________________________________
_______________________________________________________________________________
_______________________________________________________________________________
_______________________________________________________________________________
_______________________________________________________________________________
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2. Discuss the significance of sperm viability assessments.
_______________________________________________________________________________
_______________________________________________________________________________
_______________________________________________________________________________
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_______________________________________________________________________________
_______________________________________________________________________________
_______________________________________________________________________________
3. Give the clinical significance of the following result:
Result Clinical Significance
Pale yellow
coloration of semen

ST. ALEXIUS COLLEGE
Red or brownish
appearance of semen
Volume of <1.0 mL
and pH of <7.0
Decreased Motility
4. Discuss three errors that may be encountered during semen analysis that may impact the result of
the test.
Error Effect

ST. ALEXIUS COLLEGE
5. Explain Kruger’s Strict Criteria.
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_______________________________________________________________________________
6. In a 1:20 dilution, 400 sperm are counted in two WBC counting squares. Calculate the sperm
concentration per milliliter and the total sperm count in a specimen with a volume of 3 mL. Show
your solution inside the box.


Activity 9 Semenalysis

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Activity 9 Semenalysis

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ST.

Recent advancements in andrology and assisted reproductive technology (ART), coupled

Semen analysis offers a comprehensive evaluation of the male partner's reproductive

At the end of the activity, students must be able to:

Clinical Microscopy Laboratory Manual 91

Screw-capped sterile transparent plastic container

A. Patient Instructions for Sample Collection, Safe Handling and Delivery

Clinical Microscopy Laboratory Manual 92

3. Samples should be obtained by masturbation and collected in a warm (20-40ºC), sterile,

B. Information on the Sample Container and Patient Sheet

A. Semen Volume and Handling of the Specimen Container

Clinical Microscopy Laboratory Manual 93

Normal Volume: 2-5 mL

B. Semen Appearance and Odor

Clinical Microscopy Laboratory Manual 94

II. Initial Evaluation of a Wet Preparation

Clinical Microscopy Laboratory Manual 95

III. Motility Testing

Clinical Microscopy Laboratory Manual 96

b. Average proportion of duplicate slides per category

c. Difference between duplicate slides per category

G. Sperm Concentration and Sperm Count

Clinical Microscopy Laboratory Manual 97

• Only fully developed sperm should be counted.

Clinical Microscopy Laboratory Manual 98

8. Calculate the sperm concentration.

Clinical Microscopy Laboratory Manual 99

4. Air dry slides.

Normal Spermatozoa Structure

Clinical Microscopy Laboratory Manual 100

Abnormalities of Sperm Heads and Tails

18. Compute for the value of normal and abnormal forms.

𝑇𝑜𝑡𝑎𝑙 𝑁𝑜. 𝑜𝑓 𝑁𝑜𝑟𝑚𝑎𝑙 𝑜𝑟 𝐴𝑏𝑛𝑜𝑟𝑚𝑎𝑙 𝑆𝑝𝑒𝑟𝑚𝑎𝑡𝑜𝑧𝑜𝑎 𝑐𝑜𝑢𝑛𝑡𝑒𝑑

Clinical Microscopy Laboratory Manual 101

I. Calculation of Round Cells

C = round cell concentration

POST ANALYTICAL PHASE

Clinical Microscopy Laboratory Manual 102

PROCEDURE RATING COMMENTS

Clinical Microscopy Laboratory Manual 103

B. Semen Appearance and Odor

Clinical Microscopy Laboratory Manual 104

5. Count the number of spermatozoa visible in

Clinical Microscopy Laboratory Manual 105

10. Prepare a clean plain tube.

Clinical Microscopy Laboratory Manual 106

14. Add a small drop of immersion oil on the slide

Rating: 4 = Excellent 3 = Very Satisfactory 2 = Satisfactory 1 = Poor 0 = Not

Instructor: Date Evaluated:

Clinical Microscopy Laboratory Manual 107

Name: _______________________________________ Date: _________________

Clinical Microscopy Laboratory Manual 108

Clinical Microscopy Laboratory Manual 109

2. Printed pictures of the step-by-step process of semen analysis.

Clinical Microscopy Laboratory Manual 110

2. Discuss the significance of sperm viability assessments.

3. Give the clinical significance of the following result:

Result Clinical Significance

Clinical Microscopy Laboratory Manual 111

Clinical Microscopy Laboratory Manual 112

5. Explain Kruger’s Strict Criteria.

Clinical Microscopy Laboratory Manual 113

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Name: _______________________ Date: _