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Forensic Biology by Richard Li

Chapter 8: Identification of Semen. Forensic Biology by Richard Li. Biological Characteristics. Typical ejaculation 2-5 ml of semen, 160 million sperm 3 pg DNA/sperm = 480,000 ng DNA/ejaculate Only 1 ng DNA needed for STR typing! Seminal fluid Seminal vesicle fluid- 60% of ejaculate

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Forensic Biology by Richard Li

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  1. Chapter 8: Identification of Semen Forensic Biologyby Richard Li

  2. Biological Characteristics • Typical ejaculation • 2-5 ml of semen, 160 million sperm • 3 pg DNA/sperm = 480,000 ng DNA/ejaculate • Only 1 ng DNA needed for STR typing! • Seminal fluid • Seminal vesicle fluid- 60% of ejaculate • Prostatic fluid secretions- 30% of ejaculate • Acid Phospahatase (AP) & Prostate Specific Antigen (PSA) • Epididymis and bulbourethral glands- each 5% • Sperm cells- Spermatozoa

  3. Biological Characteristics • Not all semen stains contain sperm • Vasectomy- block spermatozoa • Still produces ejaculate with seminal vesicle fluid and prostatic fluid • Will not have enough sperm for DNA typing • Oligospermia- low sperm count • May or may not have enough sperm for DNA typing • Aspermia- no sperm • Will not have enough sperm for DNA typing

  4. Spermatozoa • Three distinct regions: • Head – acrosome and nucleus (with haploid DNA) • Middle Piece (mitochondria) • Tail (flagella; mobility)

  5. Acid Phosphatase • Optimal activity in acidic pH environment • Present in lysosomes • Prostate is most abundant source of AP • Half-life at 37 degrees C: 6 months • AP levels not affected by vasectomy • High levels in blood serum can be a sign of prostate cancer • See Lecture 5 for specifics of AP spot test

  6. Prostate-Specific Antigen (p30) • Major protein in seminal fluid • Also detected in urine, fecal matter, sweat, milk but in much smaller quantities • Half-life of dried stain: 3 years • Hydrolyzes semenogelins (seminal vesicle specific antigens) • Semenogelins causes semen to form gel following ejaculation • Hydrolyzing semenogelin keeps the semen fluid during ejaculation

  7. Seminal Vesicle Specific Antigen • Semenogelins • I & II • Higher concentration in seminal fluid than PSA • Not found in urine, milk, sweat • Greater specificity for semen than PSA

  8. Methods for Detecting Semen • Lighting methods (ALS) • Presumptive tests • Colorimetric • Fluroimetric • Confirmatory tests • Microscopy for spermatozoa • Antigen-antibody interactions

  9. Lighting Techniques • Locating dried stains • UV light (long-wave = “Woods Lamp”) • Argon laser • Alternate light source (ALS) • 450-495 nm • BLUEMAXX (in lab)

  10. Colorimetric Assay • AP catalyzes the removal of the phosphate group from a substrate • Positive= purple color • See Lecture 5

  11. Fluororimetric Assay • More sensitive than colorimetric • AP catalyzes the removal of the phosphate residue on the substrate 4-methylumbelliferone phosphate (MUP), which generates fluorescence under UV light • Filter paper overlay • Filter paper placed in contact with putative semen stain and then removed and taken to dark room • Sprayed with MUP • Fluorescence detected with UV lamp

  12. Confirmatory Tests • Microscopic examination • Christmas Tree Stain • Stains sperm heads red and sperm tails green Acrosomes don’t stain well in primate sperm

  13. Confirmatory Tests • Identification of prostate-specific antigen • Older methods: • Radial immunodiffusion • Rocket immunoelectrophoresis • CIE • Current method • Imunnochromatographic assay • Most sensitive • See Lecture 5 for details on PSA immunochromatography

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