Fluorescent AP Assay
Definition
A more sensitive format using 4-methylumbelliferyl phosphate (4-MUP) as substrate. AP cleaves 4-MUP to 4-methylumbelliferone, which fluoresces under UV. Useful on aged or diluted stains where colorimetric tests give equivocal results.
- Substrate
- 4-methylumbelliferyl phosphate (4-MUP)
- Product
- 4-methylumbelliferone, fluoresces under UV
- Advantage
- More sensitive than colorimetric AP tests
- Best use case
- Aged or diluted stains giving equivocal colorimetric results
Common questions
Why would an analyst switch from a colorimetric AP test to the fluorescent format mid-examination?+
If a colorimetric test on a suspected semen stain gives a weak or ambiguous colour change, often the case with old or diluted stains, the fluorescent format offers greater sensitivity by detecting fluorescence rather than a subtle colour shift, giving a clearer positive or negative result on the same type of stain.
Does a positive fluorescent AP assay confirm the stain is semen?+
No. Acid phosphatase activity is elevated in semen but is not unique to it, so a positive AP result, fluorescent or colorimetric, is a presumptive indicator that still requires a confirmatory test, such as identifying spermatozoa microscopically or a semen-specific protein or DNA marker, before semen is confirmed.
Why does acid phosphatase activity decline in aged stains, and how does the fluorescent format help there?+
AP enzyme activity degrades over time as the stain ages and is exposed to environmental conditions, weakening colorimetric colour development. The fluorescent substrate's greater sensitivity allows the assay to still detect the reduced enzyme activity remaining in an older stain.
Related terms
- Acid Phosphatase (AP)
- An enzyme present at high concentrations in seminal plasma. The Brentamine Fast Violet B (or SERI AP) spray test detects acid phosphatase...
- Brentamine Fast Violet B Test
- A two-component spot test in which sodium alpha-naphthyl phosphate (substrate) is hydrolysed by AP to alpha-naphthol, which couples with fast violet B...
- False Positive
- A test result that indicates the presence of a target analyte when it is absent. In forensic serology this may mean incorrectly...
- Prostatic Acid Phosphatase (PAP)
- The AP isoform secreted by acinar epithelial cells of the prostate gland, found in seminal plasma at concentrations of 25-1,000 U/mL, orders...
- Quantitative AP Threshold
- A numerical activity level, expressed in units per litre of extract, above which a result is treated as semen-positive. Common thresholds range...