D. Semen

Testes

Spermatogenesis

Epididymis

Sperm maturation

Testes

Epididymis

5% sperm

Seminal vesicles

Provide alkaline fluid and nutrients for sperm

Seminal vesicles

60-70% Alkaline secretion

Prostate gland

Secrete enzymes for coagulation and liquefaction

Prostate gland

20-30% Acidic secretion (citric acid, ACP, zinc)

Bulbourethral glands

Neutralize prostatic fluid and vaginal acidity

Bulbourethral glands

5% Alkaline mucus

1. Masturbation

2. Condom method

3. Vaginal aspiration

4. Coitus interruptus

Specimen collection of Semen

Masturbation

recommended method for routine semen analysis

Condom method

Silastic or nonlubricant-containing rubber or polyurethane condoms

Vaginal aspiration

for post-coital tests

Coitus interruptus

prone to incomplete collection

Patient preparation

1. sexual abstinence of 2-7 days

2. must empty bladder before collection

Specimen container

prewarmed sterile glass or plastic containers

Transport

kept at 37 °C, delivered to the laboratory within 1h of collection

Fertility testing

2-3 samples tested at 2-week intervals

2 abnormal samples considered significant

Fructose test

transported in ice & frozen within 2h of collection

grayish-white, translucent

Normal appearance of Semen

Yellow semen

Pyospermia, urine contamination, prolonged abstinence

Red/ rust color semen

Bleeding

Turbid semen

Infection (presence of WBC & bacteria)

Clear semen

Azoospermia

2-5 mL

Normal volume of semen

< 2 mL of semen

Improper functioning of one of the semen-producing organs

> 5 mL of semen

Prolonged abstinence

30 - 60 mins

Normal liquefaction of semen

> 2 hours liquefaction of semen

Deficiency in prostatic enzymes (Prostatic insufficiency)

7.2 - 8.0

Normal pH of semen

pH= < 7.2 of semen

Increased prostatic fluid

pH= > 8.0 of semen

Infection within the reproductive tract

pours in droplets (0)

Normal viscosity of semen

Clumped, stringy, or gel-like (4)

Deficiency in prostatic enzymes

1. Sperm motility

2. Sperm concentration and count

3. Sperm morphology

Routine Microscopic Examination of Semen

Sperm motility

Performed on well mixed, undiluted semen immediately upon liquefaction

Sperm motility

Speed and direction are both evaluated using approximately 20 hpfs:

Sperm motility = 4

Rapid, straight line motility (strong linear or forward progression)

Sperm motility = 3

Slower speed, some lateral movement (moderate linear or forward progression)

Sperm motility = 2

Slow progression, noticeable lateral movement (slow nonlinear or meandering progression)

Sperm motility = 1

No forward progression

Sperm motility = 0

No movement (immotile)

> 50% motile with quality > 2 within 1h

Normal motility

1. midpiece and tail abnormalities

2. male anti-sperm antibodies

Causes of abnormal motility

Sperm concentration and count

Dilution: 1:20 using sodium bicarbonate in formalin, saline, or distilled water

> 20,000,000/mL

Sperm concentration normal values

> 40,000,000 / eiaculate

Sperm count normal values

1. azoospermia

2. oligospermia

Clinical significance of abnormal counts

Sperm morphology

Evaluation is done on a thin smear stained using Wright's, Giemsa, or Papanicolaou (best stain)

Sperm morphology

200 sperm are evaluated under OlO for abnormalities in the head, midpiece, and tail

oval-shaped head

approximately 3x5 um

tail

approximately 45 um long

Semen

no big cytoplasmic droplet

acrosome

50% of the head or covers 2/3 of the nucleus

> 14%

% Normal forms for strict criteria

> 30%

% Normal forms for routine criteria

1. Decreased motility with normal count

2. Decreased sperm count

3. Decreased motility with clumping

4. Immunobead assay

5. Normal analysis with continued infertility

Abnormal Result of Semen Testing

Decreased motility with normal count

Presence of dead sperm

Viability testing using Eosin-Nigrosin stain

Work-up of decreased motility with normal count

1. bluish white

2. red

Decreased motility with normal count

1. Viable sperm

2. Dead sperm

Decreased sperm count

Lack of seminal vesicle support medium

Decreased sperm count

Fructose determination using Resorcinol test

Decreased motility with clumping

Male antisperm antibodies

Decreased motility with clumping

Incubation with male serum;

Mixed Agglutination Reaction, Immunobead assay

Normal analysis with continued infertility

1. Female antisperm antibodies

2. Failure of ovum penetration

3. Inability to penetrate partner's midcycle cervical mucus

4. Lack of membrane integrity and sperm viability

Incubation with female serum

Female antisperm antibodies work-up

1. Hamster egg penetration

2. In vitro acrosome reaction

Failure of ovum penetration work-up

Cervical mucus penetration

(Sims-Huhner test →spxipost coital vaginal aspirate)

Inability to penetrate partner's midcycle cervical mucus work-up

Hypo-osmotic swelling test

Lack of membrane integrity and sperm viability work-up

Round cells

immature sperm (spermatids) or leukocytes (primarily POD-positive neutrophils)

Microbial Testing

aerobic and anaerobic cultures

Microbial Testing

for Chlamydia trachomatis, Mycoplasma hominis, and Ureaplasma urealyticum

1. Neutral a-glucosidase

2. Zinc

3. Citric acid

4. Acid phosphatase

Chemical Testing

Neutral a-glucosidase

> 20 mU/ejaculate

Zinc

> 2.4 umol/ejaculate

Neutral a-glucosidase

for detecting disorder of the epididymis

Citric acid

≥ 52 umol/ejaculate

Acid phosphatase

≥200 units/ejaculate

1. Zinc

2. Citric acid

3. Acid phosphatase

for delayed liquefaction or prostatic insufficiency

Post-vasectomy analysis

involves examining a wet preparation using Phase-contrast Microscope for the presence of motile and nonmotile sperm

Post-vasectomy analysis

done routinely at monthly intervals beginning at 2 months post-vasectomy and continuing until 2 consecutive monthly specimens show no spermatozoa