SEMEN ANALYSIS Flashcards by Kyra Catedral

Patients with abnormal results on the routine semen analysis performed in the clinical laboratory often are referred to specialized_____ laboratories for further testing to determine the need for______

andrology

in vitro fertilization (IVF)

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In addition to fertility testing, the clinical laboratory performs________ and ________ to determine the presence of semen.

postvasectomy semen analysis and forensic analyses

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Semen is composed of four fractions that are contributed by the (5)

testes
epididymis
seminal vesicles
prostate gland
bulbourethral glands

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The_____ are paired glands in the scrotum that contain the______ for the secretion of sperm.

The external location of the scrotum contributes to a lower scrotum temperature that is optimal for sperm development.

testes

seminiferous tubules

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In the epididymis, the sperm mature and develop______.

The entire process takes approximately_____.

The sperm remain stored in the_____ until ejaculation, at which time they are propelled through the_____ to the ejaculatory ducts.

flagella

90 days

epididymis

ductus deferens (vas deferens)

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Germ cells for the production of spermatozoa are located in the epithelial cells of the_____.

Specialized______ provide support and nutrients for the germ cells as they undergo mitosis and meiosis (spermatogenesis).

When spermatogenesis is complete, the immature sperm (nonmotile) enter the______.

seminiferous tubules

Sertoli cells

epididymis

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The ejaculatory ducts receive both the__________ and ________

sperm from the ductus deferens and fluid from the seminal vesicles

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The_______ produce most of the fluid present in semen (_______%), and this fluid is the transport medium for the sperm.

The fluid contains a high concentration of____ and _____

seminal vesicles

60% to 70%

fructose and flavin.

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Spermatozoa metabolize the______ for the energy needed for the flagella to propel them through the female reproductive tract.

In the absence of it, sperm do not display motility in the semen analysis.

fructose

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_____ is responsible for the gray appearance of semen, as well as the blue to yellow fluorescence when semen is visualized under ultraviolet light (______).

Various proteins secreted by the seminal vesicles are involved in the coagulation of the ejaculate.

Flavin

Wood’s lamp

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The muscular______, located just below the blad-der, surrounds the upper urethra and aids in propelling the sperm through the urethra by contractions during ejaculation.

prostate gland

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Approximately______% of the semen volume is acidic fluid produced by the prostate gland.

20% to 30%

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The milky acidic fluid contains high concentrations of (4) responsible for both the coagulation and liquefaction of the semen after ejaculation.

Prostatic fluid

acid phosphatase, citric acid, zinc, and proteolytic enzymes

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The_______, located below the prostate, contribute about____% of the fluid volume in the form of a thick, alkaline mucus that helps neutralize acidity from the prostate secretions and the vagina.

bulbourethral glands

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It is important for semen to be_______ pH to neutralize the vaginal acidity present as a result of normal bacterial vaginal flora.

Without this neutralization, sperm motility would be diminished

alkaline

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Most of the sperm are contained in the______ portion of the ejaculate, making complete collection essential for accurate testing of both fertility and postvasectomy specimens.

first

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When a part of the first portion of the ejaculate is missing,

the sperm count will be______,
the pH will be______, and
the specimen will______.

decreased

falsely increased

not liquefy

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When part of the last portion of ejaculate is missing,

the semen volume is_____,
the sperm count is_____,
the pH is______, and
the specimen will______.

decreased

falsely increased

falsely decreased

not clot

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Specimens are collected after a period of sexual abstinence of at least _______ and not more than ______

Specimens collected after prolonged abstinence tend to have higher _____ and decreased ______

at least 2 days to not more than 7 days

higher volumes and
decreased motility

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When performing fertility testing, the World Health Organization (WHO) recommends that_____ or ____ specimens be collected not less than_____ or more than_____apart, with two abnormal specimens considered significant.

two or three

7 days; 3 weeks

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The laboratory should provide the patient with warm sterile glass or plastic containers.

Whenever possible, the specimen is collected in a room provided by the laboratory.

However, if this is not appropriate, the specimen should be kept at room temperature and delivered to the laboratory within____ of collection.

1 hour

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_______ is not a reliable means of semen collection because the first portion of the ejaculate, which contains the highest number of spermatozoa, may be lost and the low pH of the vaginal fluid may affect sperm motility.

Coitus interruptus

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Specimens awaiting analysis should be kept at_____.

Specimens should be collected by_____ into the container.

If this is not possible, only (3) condoms should be used.

Ordinary condoms are not acceptable because they contain______.

37°C

masturbation

nonspermicidial,
nonlubricant-containing rubber or
polyurethane condoms

spermicides

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Parameters reported include (6)

appearance
volume
viscosity
pH
sperm concentration and count
motility
morphology

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Normal semen has a ______color, appears______, and has a characteristic_____ odor. When the sperm concentration is very low, the specimen may appear almost_____.

gray-white translucent musty clear

Increased white turbidity indicates the presence of______ within the reproductive tract.

white blood cells (WBCs) and infection

During microscopic examination, WBCs must be differentiated from_________. The ________ reagent strip test may be useful to screen for the presence of WBCs.

immature sperm (spermatids) leukocyte esterase

Varying amounts of red coloration are associated with the presence of_______ and are abnormal. Yellow coloration may be caused by_______,_______ and _____.

red blood cells (RBCs) urine contamination; specimen collection after prolonged abstinence; and medications

is toxic to sperm, thereby affecting the evaluation of motility.

Urine

A fresh semen specimen is clotted and should liquefy within______ after collection; therefore, recording the time of collection is essential for evaluating semen liquefaction. Failure of liquefaction to occur within 60 minutes may be caused by a deficiency in ______and should be reported.

30 to 60 minutes prostatic enzymes

Analysis of the specimen cannot begin until______ has occurred.

liquefaction

If after 2 hours the specimen has not liquified, an equal volume of __________ or ________ , such as ***alpha-chymotrypsin or bromelain,*** may be added to induce liquefaction and allow the rest of the analysis to be performed.

Dulbecco's phosphate-buffered saline (DPBS) or proteolytic enzymes

Delivery of the semen specimen to the laboratory within _____of collection and documentation of the collection time are critical to evaluate liquefaction.

1 hour

Volume Normal semen volume ranges between_______. It can be measured by pouring the specimen into a clean graduated cylinder calibrated in 0.1-mL increments. Increased volume may be seen after periods of______. Decreased volume is associated more frequently with______ and may indicate improper functioning of one of the semen-producing organs, primarily the seminal vesicles. Also, incomplete specimen collection must be considered

2 and 5 mL extended abstinence infertility

Preparation of Dulbecco's Phosphate-Buffered Saline To prevent precipitation, add______ to the 1-L flask slowly, stirring continuously. Adjust the pH to 7.4 with 1 mol/L______

calcium chloride dehydrated solution sodium hydroxide (NaOH)

refers to the consistency of the fluid and may be related to specimen liquefaction.

Specimen viscosity

Droplets that form threads longer than_____ are considered ***highly viscous*** and are recorded as abnormal. Ratings of____ (watery) to___ (gel-like) can be assigned to the viscosity report. Viscosity also can be reported as ___,____,____

2 cm 0; 4 low, normal, or high.

Increased viscosity and incomplete liquefaction ***impede testing for*** (4)

sperm motility sperm concentration antisperm antibody detection measurement of biochemical markers

The pH of semen indicates the balance between the pH values from the acidic prostatic secretion and the alkaline seminal vesicles secretion. The pH should be measured within _____of ejaculation due to the loss of_____ that occurs

1 hour CO2

______pH indicates infection within the reproductive tract. A______ pH may be associated with increased prostatic fluid, obstruction of the ejaculatory duct, or poorly developed seminal vesicles.

Increased decreased

***Various factors can affect sperm concentration,*** such as (3) therefore, ***more than one semen specimen*** should be evaluated for infertility studies.

the days of sexual abstinence before the collection infection stress

Reference values for sperm concentration are commonly listed as_________ ; concentrations between ______are considered borderline.

greater than 20 to 250 million sperm per milliliter 10 and 20 million per milliliter

***The total sperm count*** for the ejaculate can be calculated by_____________

multiplying the ***sperm concentration*** by the ***specimen volume.*** (Sperm conc. X spx. volume)

Total sperm counts greater than _______per ejaculate are considered normal (20 million per milliliter X 2 mL).

40 million

Reference values for sperm concentration are commonly listed as_________ ; concentrations between ______are considered borderline.

greater than 20 to 250 million sperm per milliliter 10 and 20 million per milliliter

In the clinical laboratory, sperm concentration is usually performed using the_______. The sperm are counted in the same manner as cells in the cerebrospinal fluid cell count, that is, by diluting the specimen and counting the cells in the chamber. The amount of the dilution and the number of squares counted vary among laboratories.

Neubauer counting chamber.

Sperm concentration (Neubauer) The dilution used most commonly is______ prepared using a mechanical (positive-displacement) pipette. Dilution of the semen is essential because it immobilizes the sperm before counting. The ***traditional diluting fluid*** contains ______ and _____, which immobilize and preserve the cells; however, good results also can be achieved using_____ and ____

1:20 sodium bicarbonate and formalin saline and distilled water

Sperm concentration (Neubauer) Both sides of the hemocytometer are loaded and allowed to settle for______ mins.; then they are counted, and the counts should agree within____%. An average of the two counts is used in the calculation. If the counts do not agree, both the dilution and the counts are repeated. Counts are performed using either _______ or _____microscopy. The addition of a stain, such as_____, to the diluting fluid aids in visualization when using bright-field microscopy.

3 to 5 minutes; 10% phase or bright-field microscopy crystal violet

Sperm concentration ( Neubauer) ***Only fully developed sperm*** should be counted. Immature sperm and WBCs, often referred to as______, must not be included. However, their presence can be significant, and they may need to be identified and counted separately.

"round" cells

A count________ is associated with ***inflammation or infection of the reproductive organs*** that can lead to infertility. The presence of _______indicates ***disruption of spermatogenesis.*** This may be caused by viral infections, exposure to toxic chemicals, and genetic disorders.

greater than 1 million leukocytes per milliliter more than 1 million spermatids per milliliter

sperm concentration is reported in_________ rather than microliters.

millions per milliliter

The total sperm count is calculated by…

multiplying the number of sperm per milliliter by the specimen volume. (No. of sperm per ml/ spx. volume)

should be assessed using a well-mixed, liquefied semen specimen within 1 hour of specimen collection.

Sperm motility

Sperm motility The percentage of sperm showing actual forward movement can be estimated after evaluating approximately______. An alternative procedure is to examine______ per slide and count the percentages of the different motile categories using a manual cell counter.

20 high-power fields 200 sperm

Motility is evaluated by both_____ and ______. Grading can be done using a scale of 0 to 4, with 4 indicating ______and 0 indicating______

speed and direction rapid, straight-line movement no movement

A minimum motility of____% with a rating of 2.0 after 1 hour is considered normal.

50%

Sperm motility The WHO uses a rating scale of… Interpretation states that within 1 hour, 50% or more of the sperm should be motile in categories_____, or 25% or more should show_____

a, b, c, and d a, b, and c progressive motility (a and b).

Sperm Motility Grading ( Motile with rapid, straight-line motility) Grade WHO Criteria

4.0 a

Sperm Motility Grading ( Motile with slower speed, some lateral movement) Grade WHO Criteria

3.0 b

Sperm Motility Grading ( Motile with slow forward progres-sion, noticeable lateral movement) Grade WHO Criteria

2.0 b

Sperm Motility Grading ( Motile without forward progression) Grade WHO Criteria

1.0 c

Sperm Motility Grading ( No movement ) Grade WHO Criteria

0 d

The ***WHO Laboratory Manual for the Examination and Processing of Human Semen (2010)*** currently recommends a simpler system for grading motility that does not include speed because of the difficulty in standardized reporting. Motility is graded as (3) Motility must be specified as total motility (PM and NP) or progressive motility (PM).

progressive motility (PM) nonprogressive motility (NP) immotility (IM)

Alternative Sperm Motility Grading Criteria Sperm moving linearly or in a large circle

Progressive motility (PM)

Alternative Sperm Motility Grading Criteria Sperm moving with an absence of progression

Nonprogressive motility (NP)

Alternative Sperm Motility Grading Criteria No movement

Immotility (IM)

provides objective determination of both sperm velocity and trajectory (direction of motion). Also, sperm concentration and morphology are included in the analysis.

com-puter-assisted semen analysis (CASA)

Sperm morphology is evaluated with respect to the structure of the (4). Abnormalities in____ morphology are associated with poor ovum penetration, whereas abnormalities in the (3) affect motility.

head, neckpiece, midpiece, and tail head neckpiece, midpiece, and tail

The normal sperm has an oval-shaped head approximately______ with a long, flagellar tail approximately______

5 um long and 3 um wide 45 um long

Critical to ovum penetration is the enzyme-containing _____located at the tip of the head.

acrosomal cap

The acrosomal cap should encompass approximately____ of the head and cover approximately____ of the sperm nucleus. The_____ attaches the head to the tail and the midpiece.

half; two thirds neckpiece

The midpiece is approximately ____ long and is the thickest part of the tail because it is surrounded by a_____ that produces the energy required by the tail for motility.

7 um mitochondrial sheath

Staining for Sperm Morphology can be performed using: (4) ; the stain used is a matter of laboratory preference. Air-dried slides are stable for____

Wright's Giemsa Shorr Papanicolau’s stain 24 hours

______ can be evaluated at room temperature or at 37°C. When assessing it at 37°C, the specimen should be incubated at this temperature and the preparation made with pre-warmed slides and cover slips.

Sperm motility

Sperm Morphology At least_____ sperm should be evaluated and the percentage of abnormal sperm reported. Abnormalities in head structure that are identified routinely include (5)

200 double heads giant and amorphous heads pinheads tapered heads constricted heads

Additional parameters in evaluating sperm morphology include measuring ***head, neck, and tail size; measuring acrosome size; and evaluating for the presence of vacuoles.*** Inclusion of these parameters is referred to as______

Kruger's strict criteria.

Normal values for sperm morphology depend on the evaluation method used and vary from greater than____ normal forms when using ***routine criteria*** to greater than_____ normal forms when using ***strict criteria.***

> 30% > 14%

Additional Testing The most common are tests for: (4)

sperm vitality seminal fluid fructose level sperm agglutinins microbial infection

Abnormal Result Possible Abnormality - Decreased motility with normal count - Vitality Test???

Eosin-nigrosin stain

Abnormal Result Possible Abnormality - Decreased count - Lack of seminal vesicle support medium Test???

Fructose level

Abnormal Result Possible Abnormality - Decreased motility with clumping - Male antisperm antibodies Test (2)

Mixed agglutination reaction and im-munobead tests Sperm agglutination with male serum

Abnormal Result Possible Abnormality - Normal analysis with continued infertility - Female antisperm antibodies Test???

Sperm agglutination with female serum

Sperm vitality should be assessed _____of ejaculation. Vitality is evaluated by mixing the specimen with an _____stain, preparing a smear, and counting the number of ________ using a_______ or ______ microscope

within 1 hour eosin-nigrosin dead cells in 100 sperm bright-field or phase-contrast microscope.

_______ may be suspected when a specimen has a ***normal sperm concentration*** with ***markedly decreased motility.***

Decreased sperm vitality

Eosin-Nigrosin stain Living cells are not infiltrated by the dye and remain______, whereas dead cells stain______ against_____. Normal vitality requires_____ living cells and should correspond to the motility evaluated previously.

bluish white red against the purple background 50% or more

The presence of a large proportion of ***vital but immobile cells*** may indicate a defective______, whereas a ***high number of immotile and nonviable cells*** may indicate______ pathology

Flagellum epididymal

may be caused by lack of the support medium produced in the seminal vesicles, which can be indicated by a fructose level that is low to absent in the semen.

Low sperm concentration

are caused by abnormalities of the seminal vesicles, bilateral congenital absence of the vas deferens, obstruction of the ejaculatory duct, partial retrograde ejaculation, and androgen deficiency.

Low fructose levels

A normal quantitative level of fructose is equal to or greater than______ per ejaculate. This can be determined using spectrophotometric methods. Specimens for fructose levels should be tested within_______ or _____ to ***prevent fructolysis.***

13 umol per ejaculate. 2 hours of collection or frozen

Antisperm antibodies can be present in both men and women. They may be detected in (3) and are considered a possible cause of infertility.

semen, cervical mucosa, or serum

Specimens can be screened for the presence of fructose using the______ test that produces an_____ color when fructose is present

resorcinol test orange

Under normal conditions, the______ separates sperm from the male immune system. When this barrier is disrupted, as can occur ***after surgery, vasectomy reversal (vasovasostomy), trauma, and infection,*** the antigens on the sperm produce an immune response that damages the sperm. The damaged sperm may cause the production of antibodies in the female partner.

blood-testes barrier

The presence of______ in a male subject can be suspected when clumps of sperm are observed during a routine semen analysis. _______ cause sperm to stick to each other in a head-to-head, head-to-tail, or tail-to-tail pattern. The agglutination is graded as "few," "moder-ate," or "many" on microscopic examination.

antibodies Sperm-agglutinating antibodies

The presence of_______ indicates infection within the reproductive system, frequently the____.

more than 1 million leukocytes per millimeter prostate

Routine aerobic and anaerobic cultures and tests for (CUM3) are performed most frequently.

Chlamydia trachomatis Ureaplasma urealyticum Mycoplasma hominis

decreased fructose levels are associated with a lack of seminal fluid, decreased neutral a-glucosidase, glycerophospho-choline, and L-carnitine suggest a disorder of the_______.

epididymis

Microscopically examining the vaginal fluid specimen for the presence of sperm may be possible, with the best results being obtained by enhancing the specimen with______ and examining under_____.

xylene phase microscopy

The presence of _______in a female subject results in a normal semen analysis accompanied by continued infertility. The presence of it in women may be demonstrated by ***mixing the semen with the female cervical mucosa or serum*** and observing for agglutination. A variety of immunoassay kits are available for both semen and serum testing.

antisperm antibodies

Decreased zinc, citric acid, glutamyl transpeptidase, and acid phosphatase indicate a lack of_____

prostatic fluid

Motile sperm can be detected for up to______ after intercourse, whereas non-motile sperm can persist for_____. As the sperm die off, only the heads remain and may be present for_____ after inter-course.

24 hours 3 days 7 days

Rape ***Seminal fluid*** contains a high concentration of_______, so detecting this enzyme can aid in determining the presence of semen in a specimen. A more specific method is the detection of_______, which is ***present even in the absence of sperm.*** Often, further information can be obtained by performing______ and _____ on the specimen.

prostatic acid phosphatase seminal glycoprotein p30 (prostatic specific antigen [PSAl]) ABO blood grouping and DNA analysis

is a much less involved procedure compared with infertility analysis because the only concern is the presence or absence of spermatozoa.

Postvasectomy semen analysis

Postvasectomy Semen Analysis Specimens are tested routinely at monthly intervals, beginning at_____ postvasectomy and continuing until two consecutive monthly specimens show no spermatozoa.

2 months

Postvasectomy Semen Analysis Recommended testing includes examining a wet preparation using______ microscopy for the presence of motile and nonmotile sperm. A negative wet preparation is followed by specimen centrifugation for_____ and examination of the sediment.

phase microscopy 10 minutes