1. Chapter 8: Identification of Semen

2.  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 2

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4.  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

5.  Three distinct regions:  Head – acrosome and nucleus (with haploid DNA)  Middle Piece (mitochondria)  Tail (flagella; mobility) 5

6.  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

7.  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

8.  Semenogelins  I & II  Higher concentration in seminal fluid than PSA  Not found in urine, milk, sweat  Greater specificity for semen than PSA 8

9.  Lighting methods (ALS)  Presumptive tests  Colorimetric  Fluroimetric  Confirmatory tests  Microscopy for spermatozoa  Antigen-antibody interactions 9

10.  Locating dried stains  UV light (long-wave = “Woods Lamp”)  Argon laser  Alternate light source (ALS) ▪ 450-495 nm ▪ BLUEMAXX (in lab) 10

11.  AP catalyzes the removal of the phosphate group from a substrate  Positive= purple color  See Lecture 5 11

12.  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

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14.  Microscopic examination  Christmas Tree Stain  Stains sperm heads red and sperm tails green 14 Acrosomes don’t stain well in primate sperm

15.  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 15