Semen Reviewer (HY)
HIGH-YIELD EXAM REVIEWER — Seminal Fluid & Semen Analysis
MLS 419 | AUBF
SEMEN COMPOSITION — Quick Table
Component | % |
|---|---|
Seminal vesicle fluid | 60–70% (MOST) |
Prostatic fluid | 20–30% |
Spermatozoa | 5% |
Bulbourethral gland | 5% |
Secretions = 95% of semen. Sperm = only 5%.
STRUCTURES & THEIR KEY ROLES
Structure | Key Secretion / Role | Buzz Word |
|---|---|---|
Testes | Spermatogenesis via seminiferous tubules | "Sperm factory" |
Seminal vesicle | Fructose (energy), Flavin (color/UV), Coagulation proteins (viscosity) | "60–70%, most fluid" |
Prostate gland | Acid phosphatase, citric acid, zinc → liquefaction | "Liquefaction" |
Bulbourethral gland | Alkaline mucus → neutralizes acid | "Buffer / pH protector" |
SPERMATOGENESIS — High-Yield Flow
Mitosis (germ cells)
→ Meiosis (spermatogenesis) — Sertoli cells nurture
→ Spermatids formed (immature, NO tail, NON-motile)
→ Mature in epididymis (90 days)
→ Ejaculated via vas deferens → ejaculatory duct
Spermatids = Round Cells
Look like WBCs microscopically
↑ Round cells → infection (WBC) OR disrupted spermatogenesis (spermatid)
Differentiate using: Peroxidase stain
SPECIMEN COLLECTION — Must-Knows
Factor | Rule |
|---|---|
Abstinence | 2–7 days (>7 days = ↑ volume, ↓ motility) |
Preferred method | Masturbation |
If collected at home | Deliver within 1 hour, at room temperature |
Storage before analysis | 37°C |
Fertility testing (WHO) | 2–3 collections, 1–3 weeks apart; 2/3 abnormal = significant |
Incomplete collection consequences:
Missed part | Consequence |
|---|---|
First portion | ↓ sperm concentration, ↑ pH, failure to liquefy |
Last portion | False ↓ volume, ↓ pH, can't clot |
NOT acceptable: Ordinary condoms (spermicides), coitus interruptus (loses first portion + vaginal pH affects motility)
APPEARANCE
Normal: Gray-white, translucent, musty odor (odor is NOT reported)
Color | Cause |
|---|---|
White turbidity | WBCs / infection |
Red | RBCs |
Yellow | Urine contamination OR prolonged abstinence |
Urine is toxic to sperm → affects motility.
LIQUEFACTION
Normal: 30–60 minutes
Failure to liquefy = prostatic enzyme deficiency or incomplete collection
Cannot proceed with analysis until liquefied
If not liquefied in 2 hours, induce with:
DPBS (Dulbecco's phosphate-buffered saline)
Proteolytic enzymes (alpha-chymotrypsin or bromelain)
NORMAL VALUES — Master Table
Parameter | Normal Value |
|---|---|
Volume | 2–5 mL |
Viscosity | 0–4 scale; droplets, not stringy (WHO: threads <2 cm) |
pH | 7.2–8.0 (alkaline) |
Sperm concentration | >20–250 million/mL |
Borderline low | 10–20 million/mL |
Sperm count | >40 million/ejaculate |
Motility | >50% at grade 2.0 |
Morphology (routine) | >30% normal forms |
Morphology (Kruger's strict) | >14% normal forms |
Sperm vitality | ≥50% living |
Seminal fructose | >13 micromoles/ejaculate |
Leukocytes | <1 million/mL |
MAR test (anti-sperm Ab) | <10% sperm attached |
Immunobead test | <50% sperm with beads |
pH — Cause & Effect
pH | Cause |
|---|---|
↑ pH (>8.0) | Infection |
↓ pH (<7.2) | ↑ prostatic fluid, obstruction, poorly developed seminal vesicles |
Formula: Prostatic (acidic) + Bulbourethral (alkaline) + Seminal vesicle (alkaline) = Net alkaline pH
SPERM CONCENTRATION — Neubauer Chamber Steps
Dilute 1:20 with sodium bicarbonate + formalin (immobilizes sperm)
Load both sides of chamber
Wait 3–5 minutes to settle
Count using RBC area (5 boxes)
Count mature sperm only (with tail) — round cells excluded
Both sides must agree within 10% margin; if not → repeat
Shortcut: Average count × 1,000,000 = concentration/mL
Example: L=22, R=20 → Ave=21 → 21 million/mL × 5 mL = 105 million/ejaculate
Why is normal count >40 million? Lowest volume (2 mL) × Lowest concentration (20 M) = 40 million/ejaculate
MOTILITY GRADING
Grade | WHO | Movement |
|---|---|---|
4.0 | a | Rapid, straight-line |
3.0 | b | Slower, some lateral |
2.0 | b | Slow forward, noticeable lateral — MINIMUM NORMAL |
1.0 | c | No forward progression |
0 | d | No movement |
Normal = >50% at grade 2.0
WHO 2010 Alternative:
PM = Progressive motility (linear or large circle)
NP = Nonprogressive motility
IM = Immotility (No speed included)
SPERM MORPHOLOGY
Part | Key Feature | Abnormal Forms |
|---|---|---|
Head | Acrosomal cap = 2/3 of head → ovum penetration | Double, giant, amorphous, pinhead, tapered, constricted |
Midpiece | Mitochondrial sheet → energy for tail | Head bends backward; affects penetration + movement |
Tail | Propulsion | Double tail, coiled tail |
Stains: Wright's, Giemsa, Shorr, Papanicolaou Count: At least 200 spermatozoa
Criteria | Normal |
|---|---|
Routine | >30% normal forms |
Kruger's (ART) | >14% normal forms (stricter; includes vacuoles, acrosome size) |
SPERM VITALITY
Use when: normal concentration + decreased motility
Stain: Eosin-negrosin
Result | Interpretation |
|---|---|
Bluish-white | Living (can't take up stain) |
Red on purple background | Dead |
Normal: ≥50% living with grade 2.0 motility
Vital but immotile → defective flagellum
Many dead cells → epididymal pathology
SEMINAL FRUCTOSE
Absent/low fructose → immotile sperm (no energy)
Screening: Resorcinol test → (+) = orange-red color
Quantitative: Spectrophotometry → Normal: >13 micromoles/ejaculate
Must be done within 2 hours (or freeze to prevent fructolysis)
ANTI-SPERM ANTIBODIES
In males: Blood-testes barrier disruption → after vasectomy reversal, trauma, infection In females: Normal semenalysis but can't conceive → suspect anti-sperm antibodies
Test | Detects | Normal |
|---|---|---|
MAR (Mixed Agglutination Reaction) | IgG (screening) | <10% sperm attached |
Immunobead | IgG, IgM, IgA + location | <50% sperm with beads |
Location matters (Immunobead):
Antibody location | Effect | Can IVF help? |
|---|---|---|
Tail | Blocks movement through cervical mucosa | Yes |
Head | Blocks ovum/cervical mucosa penetration | No |
FORENSIC SEMEN ANALYSIS
Marker | Purpose |
|---|---|
Acid phosphatase | Confirms presence of semen |
PSA | Confirms semen even in absence of sperm |
Flavin (UV/Wood's lamp) | Blue-to-yellow fluorescence confirms semen |
Detection windows after intercourse:
Sperm Type | Window |
|---|---|
Motile sperm | Up to 24 hours |
Non-motile sperm | Up to 3 days |
Sperm head only | Up to 7 days |
POST-VASECTOMY ANALYSIS
Start testing: 2 months post-op, monthly intervals
Success confirmed: 2 consecutive specimens with NO spermatozoa
Method: Wet preparation → if negative: centrifuge 10 minutes → check sediment
1 motile sperm on wet prep = FAILED vasectomy
Nonmotile sperm = acceptable (cannot fertilize)
SPERM FUNCTION TESTS
Test | What it assesses |
|---|---|
Hamster egg penetration | Actual sperm penetration ability |
Cervical mucus penetration | Movement through cervical mucus |
Hypo-osmotic swelling | Membrane integrity and viability |
In vitro acrosome reaction | Enzyme release for ovum penetration |
CASA (Automation)
MT records only: volume + pH
System measures: velocity, trajectory, concentration, morphology
Three instruments: SCA (Sperm Class Analyzer), CEROS CASA, ASQA
SCA uses: digital camera + motorized heating stage + image analysis software
RAPID-FIRE BUZZWORDS
Buzz | Answer |
|---|---|
Gray-white + musty odor | Normal semen appearance |
Immotile sperm, no fructose | Seminal vesicle problem |
Failure to liquefy | Prostatic enzyme deficiency |
Acidic pH buffer | Bulbourethral gland |
Round cells = WBC look-alike | Spermatids |
Peroxidase stain | Differentiates granulocytes from spermatids |
1 motile sperm post-vasectomy | Failed vasectomy |
2 out of 3 abnormal | Significant infertility finding |
Eosin-negrosin | Sperm vitality stain |
Blue-yellow UV glow | Flavin (forensic) |
Kruger's >14% | Strict morphology for ART |
Acrosomal cap = 2/3 head | Normal head structure |
Dead cells → epididymal pathology | Sperm vitality finding |
Head antibody → IVF useless | Immunobead key point |