cover story
Semen analysis:
the test techs
love to hate
By Susan A. Rothmann, PhD, HCLD (ABB),
and Angela A. Reese, TS
CO N T I N U I N G
e D U C AT I O N
To earn CEUs, see test on page 28.
LEARNING OBJECTIVES
Upon completion of this article, the
reader will be able to:
1. Review the purpose of performing a
semen analysis.
2. Understand the problems commonly
encountered with semen analysis.
3. List and describe the different
components of the semen analysis.
4. Learn how to handle semen
specimens properly once brought to
the lab.
5. Learn which sperm-count and
motility-assessment procedures
are more reliable and less time
consuming.
6. Learn what is the best way to
determine the viability of sperm.
7. Learn what is the best way to prepare
and stain a semen smear.
8. Become aware of the different sperm
morphology classification schemes
and which ones are recommended.
9. Learn the difference between a
procedure reference and classification
reference.
10.List the four benchmarks for
semen analysis and discuss their
importance in validating the semen
analysis.
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S
emen analysis provides a snapshot of a man’s fertility
potential and is an important and critical “gateway test”
for evaluating fertility.1,2,3,4,5 As a non-invasive and
relatively inexpensive test, semen analysis often is the first test
ordered when a couple presents for a fertility work-up or when a
man is interested in permanent contraception.6,7,8,9,10 The utility of
semen analysis in assessing reproductive toxicant exposure also
makes it an important tool for environmental and occupational
health testing.11,12,13
As a major resource for semen analysis, our technical service
department often hears from technologists that they hate semen
analysis. A major factor is that semen analysis is practically the
last routine manual microscopic test in the laboratory. The lack of
reliable, affordable, tech-friendly automation certainly does add
to the test’s unpopularity. But when we delve a little deeper, we
find that technologists often have little confidence in their results
and view semen analysis as an unwelcome chore for reasons that
can be corrected easily and inexpensively.14
Here are a few of the problems that are commonly cited:
 “Long, complex, tedious, outmoded” are the words techs
use to describe their procedures. Many use equipment and
supplies not designed for semen analysis and spend a great
deal of time to get their results. For many labs, the semenanalysis procedure has been passed on through the years
without any effort to modernize or validate it.
 “Inadequate training is a huge problem.” Semen analysis is
discussed, at most, for a few hours in medical-technology
education and often is not included in clinical training. The
testing may not be performed daily, so competency and speed
are difficult to accumulate. Hardly any one has the funds
to obtain post-graduate training or time to create a training
program — and few feel qualified to teach others. Many
of the reference books and manuals on semen analysis give
conflicting or impractical advice for the lab that primarily
performs screening semen analysis.
 “We perform semen analysis, but we do not use quality controls (QC). We know we should but we just do not.” Without
the benchmark that QC gives, knowing if a test is performed
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S e m e n ana l y sis
Table 1.
What semen analysis measures
semen volume
Fluid and protein products
of male accessory glands (indirectly)
coagulation
liquefaction
consistency (aka viscosity)
concentration
motility
Spermatogenesis
sperm maturity
morphology
viability
Inflammation or infection
leukocytes
properly is very difficult. More labs (but not all) participate
in the many available proficiency-testing (PT) programs; but,
in many labs, routine CLIA-required QC is overlooked and
competency is rarely evaluated outside of a PT challenge.
General principles of semen analysis
Semen analysis is actually a panel of tests that measure testis and
accessory gland function (see Table 1), each requiring different
technology and skills.15 Each section could be the topic of its
own article; but in this summary overview, only practices that
can make testing easier or faster will be covered. Practical and
cost-effective automation is not available for semen analysis in
most clinical lab settings. Computer-assisted semen analyzers
(CASA) are very expensive and require a large investment of
time to learn and operate.16,17,18,19 Some new optical technology
is available, but its validation has not been widely reported by
independent sources, and anecdotal reports suggest that it is
unreliable in both accuracy and operation. Still, there are simple
ways to make semen analysis less of a chore, and more reliable
and useful to practicing physicians and their patients.
Semen is the product of fluids and cells from the testis and
the male accessory glands and is composed primarily of the following20,21:
 a small amount of acidic secretions from the prostate that
contain zinc, citric acid, acid phosphatase, and prostate
specific antigen (PSA);
 secretions from the ampulla and vas deferens containing
spermatozoa; and
 alkaline secretions from the seminal vesicles that comprise
most of the semen volume and contain fructose and seminogelin.
Semen should be evaluated 60 to 90 minutes after collection.
Recording both the time the sample was collected and the time the
sample analysis was initiated is essential. In some settings where
samples are transported to a central laboratory, the elapsed time
can exceed many hours. It is important that this be avoided, but if
it cannot, this delay should at least be brought to the physician’s
attention on a report. Sperm motility decreases significantly
after three hours and continues to decline over the next six to 18
hours.22,23 If delays are inevitable, samples should be kept at room
temperature, since exposure to refrigerator or body temperatures
increases the decline in motility.22,23 Because semen-analysis test
volume is often small, scheduling the testing during a single shift
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and on limited days can reduce operational expense and ensure
that properly trained technologists are available at a pre-arranged
time. Ad hoc semen-specimen “drop-off” and subsequent STAT
analysis has no place in most hospital and reference settings, and
can be very disruptive to normal lab workflow.
First, the semen is assessed macroscopically by evaluating
volume, color, consistency (often referred to inappropriately
as viscosity), and liquefaction (the change from the coagulated
to liquid state). Any obvious unpleasant odor should be noted
as it may indicate infection or excessive sample age. Routine
measurement of pH is not necessary.20 In the case of low volume
and complete lack of sperm (azoospermia), pH may give some
indication whether the problem relates to dysfunction of the
accessory glands or specimen loss during collection, but other
tests using biochemical markers are more reliable. PSA causes
proteolysis of the seminal-vesicle protein semenogelin to cause
semen to liquefy, usually within an hour, thus loss or incomplete
secretion of the first (prostatic) secretions during collection can
cause incomplete liquefaction.24,25 Samples that fail to liquefy or
have high consistency can be difficult to mix and pipet, and this
should be noted to alert the physician that test results may be
inaccurate due to unavoidable sample-handling errors.
A number of references recommend weighing the semen
sample to get the most accurate volume measurement. This
practice seems overly stringent given the variability of ejaculation
during semen collection. Clear evidence exists that for many men,
the typical practice of collecting a semen sample by masturbation
yields less volume than found during coitus.26,27,28 Thus, volume
measures, at best, are an estimate of the man’s natural semen
output and probably an underestimate. Using a 5-mL serological
pipet gives a volume measurement that is probably as reliable as
necessary with much less effort.
Mixing a semen sample is critical for accurate sperm
counts.15,29,30 The liquefied sample should be pipetted into a conical
centrifuge tube and vortexed at a medium speed for two to three
seconds twice. During pipetting, consistency can be evaluated.
If the sample leaves the pipet in drops, the consistency is normal;
if it exits as a long strand or “thread,” the consistency is high or
abnormal. At this point, some procedures recommend placing a
drop or a 10-µL aliquot of the semen on a glass slide and cover
slipping it to make a “wet preparation” that can be examined qualitatively for the presence of bacteria, round cells, agglutination,
or aggregation of sperm to other cells or debris. This extra step
can be eliminated if a sperm-counting chamber (see Figure 1) is
used. (Note: In subsequent text, the term “wet prep” refers to the
semen sample observed in a sperm-counting chamber).
Continues on page 20
Figure 1.
MLO ■ April 2007 19
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Table 2.
Comparison of sperm motility methods
Method
Ease
Accuracy
Precision
Estimate moving sperm
Difficult
Poor
Poor
Count moving sperm
Difficult
Poor
Poor
Count non-moving and immobilized sperm
Easy
Excellent
Excellent
Sperm counting
The best way to view sperm is with a
microscope equipped with a 20x phase
contrast objective. Many clinical labs do
not use phase contrast and use a 10x objective, making the procedure much more
difficult. Adding phase optics to an existing microscope is a relatively inexpensive
way to improve semen analysis.
Many labs use a hemacytometer to
count sperm. The hemacytometer was not
designed, however, for semen or sperm
counts — and using one generates a great
deal of unnecessary labor and time, not
to mention inaccurate results. The semen must be diluted, requiring duplicate
testing of the diluted sample in order to
detect dilution errors. The chamber depth
of approximately 100 µm is completely
inappropriate for a mixture of motile and
non-moving cells. Figure 1 illustrates the
problem. As time elapses, the non-moving
cells settle to the bottom of the chamber.
The moving cells continue to swim up and
down the depth of the chamber, with the
result that all of the sperm are never in the
same focal plane, making it impossible to
get an accurate count of moving cells. A
common mistake is 1) to report no moving cells when, in fact, they are just above
the plane of view, or 2) to estimate that all
are moving when the non-motile cells are
lying on the bottom of the chamber below
the plane of view.
The hemacytometer chamber and cover
slip must be thoroughly cleaned and dried
before reusing, leaving no spermicidal
residue. Sperm cells have a tendency to
adhere to glass and can contaminate future
samples if the chamber is not cleaned
thoroughly. Repeated cleaning will gradually wear down the surface, increasing
the depth of the chamber and, potentially,
leading to incorrect sperm count and calculation values. A hemacytometer that is
used regularly should be replaced every
one to two years, although many labs
use the same hemacytometer for over a
decade.
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The better choice is to use counting
chambers designed specifically for sperm
counting and to choose one that is disposable. Sperm-counting chambers have two
advantages: They do not require dilution,
eliminating the need for duplicate counts,
and they have a depth appropriate for semen (10 µm to 20 µm), which allows viewing of the motile and immotile sperm in the
same focal plane.15,31,32 Using disposable
chambers eliminates chamber cleaning,
saving labor and inconvenience while, at
the same time, providing a volumetric loading, increasing the precision of the test.
Sperm motility
Sperm-motility testing is another area
where many mistakes are made. Many
procedures are overly complicated and
more time-consuming than they need to
be. Three methods in common use are
shown in Table 2.
The most common method for performing sperm-motility analysis is estimating the percentage of motile sperm
in several microscopic fields and computing the average. Since this is almost
completely subjective, the accuracy and
Figure 2.
precision are poor. Another difficult
method requires counting both the motile
and the non-motile sperm, then calculating
the percentage of motile. If the sperm are
moving very slowly and the sample has
very few sperm, this method can produce
reasonably accurate and precise results;
but if the sperm are moving normally and
quickly, the method is almost impossible
to perform. Since sperm swim randomly,
it is difficult to tell whether a sperm at a
given point in a chamber was counted
before and then swam to a new point. Very
rapid sperm in a concentrated specimen
are virtually impossible to count.
There are many different staining
methods used, not all suited to
semen, and some technologists even
attempt to determine morphology
from unstained wet preps —
an impossible task.
An easier, more objective and reproducible method can be recommended (see
Figure 2).15,30,33 First, a small (~100 µL)
aliquot of the well-mixed, liquefied sample
is pipetted into a 1-mL snap-top microvial.
The vial is placed in a 56˚C water bath
for about five minutes to immobilize the
sperm. While this incubation proceeds,
the fresh semen sample is loaded into a
counting chamber and only the non-motile
sperm are counted. At the completion of
the incubation period, the immobilized
sample is loaded into a counting chamber
and the number of sperm (which is also
Continues on page 22
Objective determination of sperm motility
100 µL
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Figure 3.
the total count) is determined. The difference between the two — the total count
in the immobilized sample minus the
non-motile in the fresh sample — is the
number of motile sperm. From these two
numbers, calculations can be performed
to determine the sperm concentration,
percentage of motility, and total number
of sperm (concentration multiplied by
volume). Counting non-moving sperm is
easy and reproducible, within and among
technologists.
Several reference texts, including those
published by the World Health Organization (WHO)34-37 recommend that semen
analysis include a progressive motility score derived from counting motile
sperm in separate progression categories
— rapid, slow, and non-progressive
— by attempting to determine the speed
of movement. This difficult task requires
counting the number of squares the sperm
swims through during a given amount
of time using a stopwatch and an ability
to take into account many variations in
the sperm cells’ movements — almost
impossible unless the lab has a CASA
instrument.20,38 As a consequence, many
technologists simply look at the sample
and estimate the progression subjectively.
The objective motility method described
above, however, also can be used to
discern slow and non-progressive from
rapidly progressive sperm, based on the
idea that slowly moving sperm can be
counted more easily and accurately than
rapidly moving sperm. Using the method
above, for the fresh semen, one button of
a two-button tally is used to count nonprogressive and slow-swimming sperm
(those sperm that do not move more than
one square while counting across a row of
squares) and the second button is used to
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count non-motile sperm. The immobilized aliquot is analyzed as usual. The
difference between the non-motile,
slow, and non-progressive sperm in
the fresh samples and the total sperm
in the immobilized sample is the number of rapidly progressive sperm.
Other sperm-count and motility-assessment procedures are outdated and
use needless time and effort. Many
facilities, in trying to keep the semen
sample at 37˚C throughout the semen
analysis, require a heated microscrope
stage. Since short-term sperm motility
in semen is not noticeably affected by
changes of temperature between room
air and body temperature, this practice
is unnecessary.20,22,23 Labs that measure
sperm motility at multiple times after
collection engage in a completely useless
and time-wasting measure that yields no
clinical information. Some technologists
use Gram stains to detect bacteria, but they
are usually evident during microscopic
exam of the fresh semen or in stained
smears. There is little clinical association
of bacteria with infection, and they are
probably a result of contamination during
collection. A peroxidase test can be used to
detect white blood
cells, but a tech- Figure 4.
nologist trained in
hematology smears
will rarely, if ever,
confuse a neutrophil with an immature sperm precursor cell using a
well-stained semen
smear.
counterstain.39,40,41 The method is quick
and easy to evaluate. Sperm that exclude
the stain are alive, and those that take up
the eosin are dead. Both can be visualized well against the blue-black nigrosin
counterstain (see Figure 3). In a freshly
collected sample, the percentage of motile and viable sperm should be similar,
making viability a good check of motility. Since dead sperm do not swim, the
number of viable sperm should always
be higher than or close to the number of
motile sperm. Some accrediting organizations require viability testing when sperm
motility is low in order to rule out necrozoospermia. In practical use, the threshold should be very low (less than 10% to
30% motile), since necrozoospermia as
a clinical condition is rare. Probably the
most common cause is contamination of
the sample with lubricants, most of which
are spermicidal.42,43
Sperm morphology
Sperm morphology is probably the most
confusing and time-consuming area of
semen analysis.20,44,45,46,47 There are many
different staining methods used, not all
suited to semen, and some technologists
Sperm viability
Sperm-viability
testing typically
uses a nuclearexclusion stain to
determine whether
non-motile sperm
are alive and not
able to move,
or actually dead
(necrozoospermia). Viability
testing requires a
very simple twostep staining procedure, using eosin-Y as the stain
and nigrosin as a
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even attempt to determine morphology
from unstained wet preps — an impossible task. There are many classification
systems in use, each with its own criteria
for what constitutes a normal cell. Some
systems report the location of the defects,
some report the specific types of defects,
and some only report that the cell is abnormal with no indication of why. It is
no wonder that people are uncertain what
classification or stain to use and how to
report the results.
The first step in sperm morphology is
to make a good, even semen smear — not
too thin and not too thick. Too thin and
there will not be enough sperm present
for a good evaluation. Too thick and the
sperm can be layered and difficult to focus
on clearly. Roughly made smears can also
separate the sperm heads from the tails as
an artifact, leading to an incorrectly high
percentage of midpiece abnormalities.
To make the smear, a small drop of
semen (approximately 10 μL) should be
placed near the labeled end of the slide.30
Another glass slide held at 45˚ angle is
then used to make a push smear or pull
smear. This angle can be increased or de-
Table 3.
Normal Sperm Morphology
from Common Sperm Classifications
ASCP
Normal Reference Range
MACLEOD
STRICT
WHO 4TH
WHO 2ND
WHO 3RD
>80%
>60%
>14%
>50%
>30%
shape
oval
oval
oval
smooth
border
oval
oval
acrosome
1/2 to 2/3
of head
surface*
40-70% of
head
surface
>1/3 of
head
surface
40-70% of head
surface
size
4-5 µ long
2-3 µ wide
3-5 µ long
2-3 µ wide
3-5 µ long
2-3 wide
(width = 1/22/3 length)
4-5.5 µ long
2.5-3.5 µ wide
length/width
=1.5-1.75
vacuoles
not clear
HEAD
<20% of
head area
not stated
up to 4
not
considered
slender,
straight,
regular
outline
straight,
regular
outline
straight,
regular
outline
axially
attached
axially
attached
axially
attached
<1 µ wide
length 1.5X
head
<1/3 head
width
7-8 µ long
<1/3 head
width
7-8 µ long
MIDPIECE
shape
size
1 µ wide
5 µ long
cytoplasmic droplet
considered
to be immature sperm
<1/2 of
head
area
not
considered
uniform
size
uncoiled
<1/3 of head
area
TAIL
width
1 µ at base
0.1 µ at tip
thinner
than
midpiece
length
50-55 µ long
10 x head
slender
uncoiled
regular
outline
slender
uncoiled
regular outline
> 45 µ long
> 45 µ long
creased to make the smear slightly thicker
or thinner, depending on the concentration
of the sample. Ideally, the smear should
immediately be fixed using a spray cytology fixative, dried thoroughly, and then
stored in a dry, dark area until stained.
The best stain for semen smears and
sperm is a modified Papanicolaou (Pap)
stain.34-37,48 This provides good clarity for
viewing and color differentiation between
the regions of the cell. Pap stain is also
stable over time, allowing smears to be
stored for later review. The typical Pap
staining method is time-consuming to
prepare and perform; but with many of the
stains available commercially, the results
are worth the effort.49 A three-step quick
Pap stain also can be purchased. Because
semen analysis often is performed in body
fluid sections of hematopathology, stains
such Wright-Giemsa are used. Although
rapid and available commercially in onestep and three-step kits, these stains do
not provide the same clarity and color
variation as the Pap stain (the one-step kits
should always be avoided). A particular
problem is that semen, which is almost
unnoticeable in a Pap-stained smear, stains
vivid purple with Wright-Giemsa stains,
obscuring much of the fine detail of the
sperm morphology. A third option for
semen smears is to use pre-stained blood
film slides, but the visual quality of these
for sperm cells is very poor and the stain
is not stable, thus the smears cannot be
stored for later review.
No matter what stain is used, the smear
should be evaluated using a 100x oil objective with a 10x eyepiece. Basically, each
sperm is evaluated, looking at the head,
midpiece, and tail (see Figure 4). If any of
the three major structures is abnormal, the
sperm is classified as abnormal.36 A single
key tally is used in one hand to keep track
of the number of sperm analyzed, while a
multikey tally is used with the other hand
to tally normal, borderline abnormal, or
abnormal.49 Depending on what information the ordering physician wants, some
labs will need to provide the location of
the abnormality as well (head, midpiece,
tail), and some will need to provide more
detail of the type of abnormality, such as
shape of head, size, and so forth.48,49,50,51 At
least 200 cells must be evaluated.
The major difficulty in morphology is
identifying normal sperm as determined
by which classification system is used.
Ideally, a classification scheme provides
Continues on page 24
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MLO ■ April 2007 23
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Figure 5.
Examples of normal and abnormal sperm
scheme. The Sperm Confirm Atlas of
Sperm Morphology 48 compares classification schemes and lists appropriate
references.
Benchmarks for semen analysis:
training, quality control, proficiency,
and competency testing
a standardized system that allows many
observers to compare results and should
have clinical relevance and relate in a
meaningful way to fertility.45 Over the last
50 years, five major published classification systems have endured in wide clinical practice: MacLeod,3,51 WHO Manual
2nd ed.,35 WHO Manual 3rd ed.,36 ASCP,52
and Strict, described by Menkveld53,54 and
Kruger,55,56 and promoted in the WHO
Manual 4th ed.37 Table 3 summarizes the
main components of these five schemes
in common use.
Of these, the WHO 3rd and the Strict/
WHO 4th are the modern classifications
that are most recommended by fertility
physicians. Unfortunately, there are so
many variations of the two schemes that
learning either one or comparing the
results is very difficult.46 Judging from
their individual atlases,54,56 the two main
proponents of the Strict classification do
not agree about what is a normal sperm.
Examples of basic sperm morphology
are shown in Figure 5. Much of the classification difficulty and controversy arises
over just how perfect a sperm must be to
be considered normal. In actual practice,
smearing, fixation, air-drying, and staining
can induce artifacts that must be identified
and distinguished from the sperm’s natural
morphology.
A comprehensive discussion of morphology classification is beyond the
scope of this article and can be found
elsewhere.48 To simplify the topic, we
make several recommendations that can be
implemented by most labs. For the general
reference laboratory that performs semen
analysis primarily as a screening tool, the
WHO 3rd scheme provides an appropriate
starting point for fertility evaluations.1,57
In specialized fertility testing and treatment settings, the Strict/WHO 4th criteria
is generally used.1,55,58 Keeping in mind
that neither has a clear reference standard,
a way to distinguish these two schemes
is based on classification of borderline
abnormal forms (see Table 4).
No matter which classification scheme
is used, it is vital to make sure the reference materials are correct. A very common error in morphology classification
is to confuse the procedure reference for
the classification reference. We asked
participants in our July 2006 Proficiency
Challenge and in a separate morphology class to list both their classification
system and the reference atlas they used
(see Table 5). We found that one-third of
the proficiency challenge participants and
one-fifth of the class attendees were using
the wrong atlas for their system, leading
to incorrect results. For example, if sperm
are classified using
Table 4.
the WHO Manual
4th ed. as a proceComparison of sperm classification methods
dure reference, but
cite Atlas of Sperm
WHO 3rd
Strict/WHO 4th
Morphology writReference value for normal
Over 30%
Over 14%
ten by Adelman
and Cahill52 as the
Classification of borderline abnormal Normal
Abnormal
reference atlas, the
Screening,
system actually
Utility of classification
Fertility treatment
fertility treatment
used is the ASCP
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Formal semen-analysis training is rare;
and without adequate education, it is very
difficult to feel confident about clinical
test results. Some professional societies
and groups offer instruction, but many
reference labs do not have budgets for
training when travel is involved. Selfpaced training courses15,49 and video30
are available, however, for the lab that
wants to improve the technical skills of
its staff.
Another reason that semen analysis
is the test techs love to hate is that QC is
often not used routinely. Many techs (or
their supervisors and managers) seem to
think that semen analysis is excluded from
the CLIA regulations. On the contrary,
semen analysis is a high-complexity test
requiring two levels of quality controls on
each day of patient testing.59,60 This not
only documents the ability of the technologist to perform the test correctly but also
gives him confidence in the results of the
test.61 Without a benchmark that QC gives,
knowing if a test is performed properly is
very difficult.
Adding phase optics to an
existing microscope is a relatively
inexpensive way to improve
semen analysis.
A quality control for sperm count is
like that for any other test: one that can
be analyzed in any counting chamber
or semen analyzer, in the same way as a
patient sample. In order to truly mimic a
clinical test material, the quality control
must resemble the sample being analyzed. In the case of semen analysis, no
surrogate materials come close to looking
as complex as sperm cells in semen (see
Figure 6). Sperm cells are oval and have
tails that often overlap or coil around
the head, changing the appearance and
making them harder to differentiate from
non-sperm cells. Semen contains debris,
immature germ cells, white blood cells,
and other non-sperm cells, some of which
can be confused with sperm. Although
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s e m e n ana l y sis
Table 5.
Classification system and atlas discordance
% using inappropriate atlas
% listing no reference
PT
32%
6%
Morphology class
19%
28%
latex beads are advertised as a control for
sperm count, beads are really not a valid
control.62 Latex beads are spheres, and are
easy to count since they all look the same,
and there are no other particles present to
confuse the technologist. A suspension
of human sperm is the only valid quality
control for sperm count.
Sperm-motility quality controls are
available in two formats: frozen aliquots
of a fresh sample to be thawed for analysis, and video recordings on CD-ROM or
other format. The frozen aliquots allow
the controls to be analyzed in a counting
chamber, but due to the variation in spermcell death during thawing, the motility of
frozen-thawed sperm is highly variable.
These controls also require specialized
liquid-nitrogen storage and a regular supply of liquid nitrogen, which greatly adds
to the expense of the product and to the
difficulty in safe handling of the ultra-lowtemperature materials.
Video recordings, on the other hand, are
easy to use and inexpensive. When video
of both the fresh and immobilized samples
are provided, these quality controls can be
used with any method of motility analysis.
The only equipment needed is generally a
computer and tally counter.
Quality controls for sperm morphology
must be assayed for the classification system being used. Most of the morphology
quality controls currently available are
assayed for WHO 3rd or for Strict/WHO 4th
classification schemes. QC for the Strict/
WHO 4th scheme is especially important
as viewers tend to get stricter with time
until no normal sperm can be identified.63
Assayed ranges for the older classification systems, such as ASCP, McLeod, and
WHO 2nd ed., are not commonly provided
with controls. Facilities that use one of
these classification systems must establish
control ranges by analyzing the control
slides at least 20 times.64,65 Switching to
a modern system probably makes more
sense. Morphology quality-control smears
are usually stained. If the manufacturer
uses a different stain, unstained smears
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from an assayed lot can be obtained to
stain in-house.
CLIA requires proficiency testing twice
a year for semen analysis59,60 and a number
of providers offer appropriate PT material. Labs that use automated analysis for
any part of semen analysis must be able
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MLO ■ April 2007 25
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to analyze PT materials. Proficiency testing for semen analysis
identifies many of the analytic problems discussed above. For
example, morphology proficiency-testing results often show a
range of normal from 0% to 100%.46 Providers that have innovative virtual challenges for morphology can give a great deal of
educational information about sperm classification (e.g., American
Proficiency Institute, Wisconsin State Laboratory of Hygiene).
Annual competency testing of each technologist is also required by CLIA.59 Proficiency-testing results can be used as part
of competency testing to show how well the technologist performs
compared to the peer group. Additional competency testing should
include written examination of important aspects of the topic as
well as observation of test performance by a supervisor. With so
many supervisors feeling uncertain about how semen analysis
should be performed, many labs skip over competency training
for this area of lab medicine. Online and written products are
available for documenting training and competency.
Summary
At the heart of better semen analysis is professionalism. The
walls of many labs are covered with slogans like, “at the end
of every test is a worried patient who needs an answer.” Semen
analysis is no different. At its end is a couple desperate to have
a child or start a family. In spite of the importance of semen
analysis in fertility diagnosis and treatment, it remains in most
clinical laboratories “the neglected laboratory test.”6
The tips and recommendations in this article should help any lab
improve the quality of semen analysis while reducing the effort required to produce better results. Knowledge and simple, repeatable
procedures, combined with QC and competency benchmarks, can
put the interest and satisfaction back into a test that is the gateway
for fertility treatment. After all, what is not to love about a cell that
swims and comes in so many interesting shapes? l
Acknowledgement: The authors thank Jeremy Paul for his assistance with the manuscript.
Panbio's West Nile Virus ELISA Range
Susan A. Rothmann, PhD, HCLD(ABB), is president and laboratory director of Fertility
Solutions, a company that manufactures and distributes semen-analysis quality controls,
reagents, and training materials, in Cleveland, OH. She can be reached at srothmann@
fertilitysolutions.com. Angela A. Reese, TS, is the technical service manager of Fertility
Solutions. She received a BS in biology from John Carroll University, and is certified as a
technical supervisor in andrology. She has helped to develop new semen-analysis training
tools and quality controls, particularly for sperm morphology and motility.
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