Presumptive Tests for Semen
Semen contains acid phosphatase (AP), a common enzyme in nature that occurs at a very high level in semen. This allows for a test, with a high AP threshold, to presumptively identify semen. The most common test is Brentamine Fast Blue B applied to the sample on an alpha-naphthyl phosphate substrate (see “In More Detail: Presumptive Test for Acid Phosphatase”). Brentamine Fast Blue B is a known carcinogen and must be handled accordingly. A piece of filter paper or cotton swab is moistened with sterile water and applied to the questioned stain. The reagent is added, and if an intense purple color is seen, the test is positive; if no color reaction occurs within 2 min, the test is negative. If a sample is known or suspected to be “old,” a negative result should be cautiously interpreted because AP becomes less active over time. Positive results are presumptive, remember, because many biological fluids, including vaginal secretions, contain some amount of AP. Commercial test kits are available for semen.
FIGURE 1 Between 100 and 150 million spermatozoa are found per milliliter of ejaculate. Semen (or, by extension, sperm) is considered presumptive evidence of sexual contact.
Semen can be visualized by alternative light sources at 450 nm and viewed with amber goggles. The method is useful because it works on light or dark surfaces, covers large areas, and is quick; the disadvantage is that some other fluids will also fluoresce, such as saliva and urine. Semen exhibits a blue-white fluorescence, so this test can help narrow down which stains to test for AP. Just because no fluorescence occurs, however, it cannot be assumed that no semen is present.
