Semen: Composition, Biology, Forensic Analysis
Mission, Vision, Core Values
• Mission: “A nurturing ground for an individual’s holistic development to make effective contribution to the society in a dynamic environment.”
• Vision: “Excellence and Service.”
• Core Values: Faith in God | Moral Uprightness | Love of Fellow Beings | Social Responsibility | Pursuit of Excellence.
Overview of Semen
• Definition: Viscous, whitish‐grey fluid expelled from male reproductive tract during ejaculation.
• Composition: Mixture of mature spermatozoa + seminal plasma (secretions of accessory sex glands).
• Typical ejaculation: containing
Cellular vs. Acellular Components
• Cellular
– Mature spermatozoa.
– Epithelial cells of urogenital tract.
– “Round cells”: leukocytes & immature germ cells.
• Acellular (glandular secretions)
– Testis/Epididymis: GPC, Neutral (\alpha)-glucosidase, free L-carnitine.
– Seminal vesicle: Fructose, Semenogelin, Prostaglandins.
– Prostate: Prostate Specific Antigen (PSA), , citrate.
– Cowper’s (bulbourethral) gland: Mucin.
Male Reproductive Anatomy (Key Structures)
• Testes → Epididymis → Ductus deferens → Ejaculatory duct → Prostatic urethra → Penile urethra.
• Accessory glands: 2 seminal vesicles (posterior to bladder), single prostate (inferior to bladder, around urethra), 2 bulbourethral glands (either side of membranous urethra).
Testes and Spermatogenesis
• Testes: Ovoid, long, inside scrotum; homologous to ovaries.
– LH → Leydig cells → testosterone.
– FSH → Sertoli cells → support spermatogenesis.
• Seminiferous tubule hierarchy
– Spermatogonia → spermatocytes → Meiosis I → spermatocytes → Meiosis II → spermatids → spermiogenesis → spermatozoa.
• Timing: months for production + maturation; storage in epididymal tail.
Spermatozoon Structure
• Head: Nucleus (23 chromosomes) + acrosome (digestive enzymes).
• Midpiece: Mitochondria‐rich collar for ATP generation.
• Tail (flagellum): Propulsion; progressive motility required.
Accessory Sex Glands
• Seminal Vesicles
– Paired, ~ of seminal volume.
– Secrete viscous, fructose‐rich, prostaglandin‐rich fluid; androgen dependent.
• Prostate
– Walnut sized; 30–50 concentric tubuloalveolar glands.
– Contributes of semen; doubles in size during puberty, enlarges after (usually benign).
– Secretion: slightly acidic mucin & citric acid (nutrient), PSA (liquefaction enzyme).
• Bulbourethral (Cowper’s) Glands
– Produce mucin for lubrication (~ of volume).
Sperm Morphology Abnormalities & Clinical Clues
• Macrocephaly: Giant head, aneuploidy; fertilization failure.
• Microcephaly: Small head/acrosome defect; reduced DNA.
• Pinhead: Almost DNA‐less; can indicate diabetes.
• Tapered (“cigar”) head: Linked to varicocele or chronic scrotal heat.
• Thin/narrow head: Rare; may reflect DNA breaks or varicocele.
• Globozoospermia: Round head with absent acrosome; failure to activate egg.
• Decapitated (headless) sperm: Acephalic syndrome; no genetic content.
• Tail‐less (acaudate): Seen in necrotic conditions.
Semen Analysis – Normal Reference Values (WHO-like)
• Volume:
• Sperm concentration: Total
• Motility: Total ; progressive
• Morphologically normal forms:
• Vitality: live.
• WBC: <1\times10^{6}/\text{mL}.
Descriptive Properties of Normal Semen
• Appearance: Grey-opalescent; coagulates then liquefies within 1 h.
• pH: (low pH
Biological/Forensic Characters
• Fractional contributions: Seminal vesicle , prostate , epididymis + bulbourethral each.
• Flavin content → UV fluorescence (crime-scene search).
• Vasectomy: Removes ductus deferens segment → ejaculate lacks sperm but retains accessory fluids.
• Oligospermia: Low sperm (<). Variants: Severe (<); Azoospermia (zero).
• Even in azoospermia, epithelial DNA may be recovered from fluid.
Key Forensic Enzymes & Proteins
• Acid Phosphatase (AP)
– Optimum acidic pH; prostate isoform dominates semen.
– Not reduced by vasectomy.
– Half-life: months @ (dry); shorter in wet.
– Detectable up to yr @
• Prostate Specific Antigen (PSA / P30)
– in semen; serine protease.
– Encoded by KLK3 (chromosome 19); hydrolyzes semenogelin → liquefaction.
– Found (lower levels) in urine, sweat, milk; elevated serum PSA in prostate cancer, BPH, prostatitis.
– Dry stain half-life years (room temp); wet storage rapidly degrades.
• Seminal Vesicle Specific Antigens (SVSA)
– Semenogelin I & II (SgI/SgII); high abundance; absent from urine/sweat/milk → higher specificity vs PSA.
– Form initial coagulum; degraded by PSA.
Analytical Techniques for Semen Identification
• Visual/Lighting Survey (Presumptive)
– Alternate Light Sources (ALS): Excite , view through orange goggles; semen fluoresces strongly but saliva/urine also fluoresce.
– Observational descriptors: Thick yellowish-white; starchy texture; stains appear reddish-brown/yellow/grey on fabric.
Presumptive Chemical Tests – Acid Phosphatase
• Colorimetric ((\alpha)-naphthyl phosphate + Fast Blue B)
– Transfer sample with moistened swab; add substrate + dye.
– Purple within <1 min → positive (prostatic AP).
– Tartrate inhibition or substrate choice ((\alpha)-naphthylphosphate vs phenyl phosphate) ↑ specificity.
– Reagent formulation (Sodium (\alpha)-naphthyl phosphate method):
Buffer ((\text{Acetic acid} + \text{Na acetate})) + Step-1 & Step-2 reagents (see transcript).
• Fluorometric (4-Methylumbelliferone Phosphate, MUP)
– Filter paper blot mapping → spray MUP → UV illumination → fluorescing spots denote AP activity.
Confirmatory Tests
• Microscopy for Spermatozoa
– Sample extraction → slide mount → air/heat dry.
– Christmas Tree Stain: Nuclear Fast Red (nuclei/acrosome = red) + Picroindigocarmine (midpiece/tail = green).
– SPERM HY-LITER fluorescence or Laser Capture Microdissection (LCM) enhances recovery for DNA.
• Ammonium Oxalate‐Crystal Violet (Gram-like) staining
– Reagent #1 (Crystal violet + ammonium oxalate), #2 Gram’s iodine, #3 95 % ethanol, #4 Safranin.
– Positive proteins → violet; negatives → red.
• Immunoassays for PSA (p30) or SVSA (Sg): Highly sensitive and confirmatory.
• RNA-based assays (not detailed) complement protein detection.
Clinical / Ethical / Practical Implications
• Abnormal morphology or motility → male infertility; associations: varicocele, diabetes, heat exposure.
• PSA/Sg detection pivotal in sexual assault investigations (linking perpetrator).
• Vasectomy: Socio-ethical choice for contraception; forensic consequence – sperm absence.
• Elevated serum PSA as cancer biomarker but confounded by BPH/prostatitis → informed clinical decisions.
Numerical Tables & Data Highlights
• Semen category statistics (Normospermia ⇢ Azoospermia) → correlations: decreased density, motility, morphology with severity.
• WHO normal vs oligospermia/severe categories: see values above for quick recall.
Storage & Stability Guidelines
• AP: dry @ months; wet storage accelerates loss.
• PSA: dry stain years; wet → rapid degradation.
• Best practice: Air-dry evidence, refrigerate or freeze ((\le -20^{\circ}\text{C})).
Key Connections / Integrative Notes
• Spermatogenesis relies on endocrine axis (LH/FSH/Testosterone) paralleling ovarian folliculogenesis concepts.
• Semenogelin‐PSA cascade = seminal equivalent of fibrinogen‐plasmin in blood coagulation-fibrinolysis paradigm.
• Fluorescence search parallels use of ALS for saliva/urine/blood latent detection; specificity always requires chemical or microscopic confirmation.