Fluoresence of semen (forensics geekery)

After hearing in the news that a blacklight can be used at a motel to check for semen stains on sheets and carpets, I began experimenting with that concept to see if it is at all useful (ok, I’ll wait for you to stop laughing… This is science, people!).

TL;DR: What I found is that a typical blacklight is entirely useless for detecting semen, but does cause urine, cat vomit, some soda pop, some drugs, and soap remnants to fluoresce. Several fabrics and papers are infused with whiteners from detergent or mixed into the fibers, and these will also fluoresce brightly. Liquids spilled onto such fabrics or papers may draw out these whiteners and seem to fluoresce, but it is a secondary effect rather than the liquid itself. Key point: UV is only useful to help find droplets during an investigation. It doesn’t identify what the substance is.

Semen will very slightly fluoresce blue under some lights, and orange under other light. Wet semen does not seem to fluoresce at all, it must have dried. (1) Someone wrote to me and suggested that Luminol is needed to cause dried semen to fluoresce. However, none of the literature I read indicated that Luminol is useful for anything but detecting blood, and none mentioned it being used to detect semen.

A bit of background on fluorescence
Our eyes can seen the “visible spectrum” of light, from about 400nm (nanometers) to 700nm. The typical blacklight has a predominantly ultraviolet (UV) wavelength of about 360-370nm, which is also called “longwave” ultraviolet. The bulbs are usually coated with a dark blue filter to keep most visible light from emitting, though there is always a dark violet glow. This coating leads to the industry term of “blacklight blue” or BLB. The non-coated bulbs (BL) emit a bright blue glow in addition to the UV and are used more for bug-zappers and for killing bacteria.

Fluorescence of a material is based on the wavelengths of light it absorbs, and the wavelengths of light it emits after absorbing light. We tend to notice fluorescence most when the source light is low in the visible spectrum (such as a black light), coupled with fluorescent material that emits strongly in the visible spectrum (like highlighter pens and blacklight posters). Most fluorescent minerals require an ultraviolet source wavelength that is “short” (250nm to 300nm), which is dangerous to our eyes and skin, and requires the use of UV filter glasses to safely use. Minerals will typically not fluoresce at all under a standard blacklight. The total UV light spectrum starts at the extreme end of 10nm wavelength and stops around 400nm.

The source light wavelength that causes fluorescence of semen is reported to be 415-490nm, which is not UV at all, but is in the visible violet-blue-dark green part of the spectrum. The best wavelength was 415nm according to one source (1), but another claimed that 570nm (light green) was useful without any goggles (3). The fluorescent emission peaks of semen are reported to be around 460-520 nm, though one older paper specifically mentions 622nm emission when excited with 488nm (2). Blue LEDs are sold in the 450 to 480nm range, and filters are also sold for that range. Some forensics websites are now selling blue LED flashlights for this purpose. But because there is so much visible light produced by these lights, they recommend using dark orange glasses to filter out the blue light and let the fluorescence come through.

My approach
Since the light sources used in forensics are priced so highly (due to selling to law enforcement, and the intense mercury vapor or argon laser light source), I began to experiment with lower cost solutions. Perhaps I could substitute “blu-blocker” sunglasses and use a light source I already had. The GoLite P2 for winter light therapy is said to put out about a 470nm blue light which is in the right range for a source light.

I did a lot of searching online and found a cheap blue-LED flashlight in the 455-460nm range (LED Wholesalers B001TIEHO2), and some dark orange safety glasses (UVEX S0360X) on Amazon. (insert pic) The dark orange glasses filter out the visible blue light from the flashlight, but allow the fluorescent emission of certain materials to pass through. I also bought a 56-inch square piece of orange colored transparent vinyl, with the idea that I could make it as dark orange as I like. However, though it is “transparent”, it is not optically clear so it isn’t that great for seeing clearly.
blue and orange - amazon

Results of the blue/orange approach
With this combination, I found that I could still see urine and soap (and cat vomit traces on the rug) very brightly and with good contrast. But semen still was nearly invisible. I still need to try mucus and blood.



I have no way to verify if the LEDs on my flashlight are actually in the correct spectrum, and am relying on the manufacturer’s description to be accurate.

Another issue is that the orange UVEX glasses themselves fluoresce, causing considerable fogging when the blue light is reflected off a white surface. Not sure if it is the plastic or the orange coloring that does this. A vinyl sheet that is clear didn’t fluoresce, but the orange vinyl did, so I’m assuming that it is the orange coloring that is used for plastics that is fluorescing rather than the plastic.

The Green Light approach (3)
My next experiment was to use a light-green filter over my regular CREE LED flashlight, to limit the output to the 570nm range.

I used a $5 filter advertised as a 570nm green filter, purchased from an online electronics company.

This approach is supposed to be used without goggles. I cut a square of aluminum foil and put a hole in the middle, and used clear tape to attach it. I placed this over the front of an Olight M22 Warrior flashlight on maximum output.

Well, that was disappointing. No fluorescence, no distinguishing characteristics at all for samples on wood and plastic. I didn’t pay to read the whole paper on this claim, so perhaps I missed something. I kinda doubt it though. Most examples of semen fluorescence are minimal even with advanced light filtration.

My conclusion is that semen is not particularly fluorescent.

You will definitely NOT find semen using a regular blacklight. A blacklight is useful for finding traces of soda-pop, urine, vomit, and several items listed below. The blue-light with orange goggles is the best overall for finding urine and such, as it really pops out brightly.

Remember that in the forensics field, fluorescence only helps locate samples of semen, not to identify it as a particular substance. Identification is the realm of chemical and microscopic analysis. In the quote below, it is said to be highly presumptive to do otherwise. This is because so many kinds of stains (soda-pop, urine, soap, vomit, and lots more) will all fluoresce, and far more brightly than semen.

Other things that glow brightly under a regular blacklight:

    • Tonic water (quinine makes it glow bluish) and several soft drinks with bright colors.
    • The many zillions of brightly dyed candies, and spit/drool containing dyes from pop or candy.
    • Scorpions (good way to locate them at night) and some other arthropods
    • Chalk
    • Many plastics
    • paper dust, certain kinds of cloth dust
    • According to one website, some narcotics[4] (again, only an way to locate, not a positive ID):

Cocaine (UV flashlights are used to detect this in the nostril region)
MDMA tablets (including some but not all Ecstasy tablets)

Cocaine – Cocaine having a purity of at least 87 % fluoresces clearly when illuminated with UV light.

Amphetamine – Some amphetamine having a purity of 78 % are clearly fluorescent when illuminated with UV light. Even small amounts of amphetamine are easy detectable because of their fluorescent nature.

MDMA tablets – Some MDMA tablets (i.e. Ecstasy with four-leaf clover logotypes) are clearly fluorescent with UV. Even small fragments are easily visible as they fluoresce intensely.

…semen will fluorescence due to the presence of molecules such as Flavin and Choline-conjugated proteins. The color of this fluorescence will vary from blue to yellow, depending on the light equipment used. There are many molecules (natural and artificial) that will fluoresce in a similar way as semen, and therefore, this detection technique is highly presumptive. Furthermore, not all semen stains will fluoresce under such specialized lights. Exposure of the sample to factors such as heat, humidity, oxidizing agents, and microorganisms such as bacteria and mold can affect this fluorescent activity. Semen fluorescence can also be masked by certain types of fabrics and fabric treatments. Hilton J. Kobux, D.Phil., Edmund Silenieks, and Jordana Scharnberg, B.Sc., Improving the Effectiveness of Fluorescence for the Detection of Semen Stains on Fabrics, 47 Journal of Forensic Sciences 4 (2002); S. Marshall, A. Bennett, and Dr. H. Fraval, Locating Semen on Live Skin Using Visible Fluorescence, Rofin Australia Pty Ltd. (2001).

(1) http://www.bvda.com/EN/prdctinf/semen_fluo.html
…if you illuminate dry semen with a band of light around 350 nm HPBW 40 nm (ie 330-370 nm), which is invisible to the human eye, then the semen will fluoresce, into the blue visible region (ref trace 2). The advantage of this is that you can make invisible semen stains appear visible to the naked eye…
there is an alternative to using UV when searching for semen stains. Illuminating dried semen with a band of light around 450 nm HPBW 40 nm (ie 430-470 nm) will produce strong orange fluorescence (ref trace 3) in a broad region with a maximum around 520 nm.

Best results were found with 415nm excitation (source light) and a 475nm high pass filter. Light appears greenish, semen shows up bluish.
2nd best was 450nm excitation with 590nm high pass filter. Light appears orange, semen appears yellow-orange.
Source light in both cases was Polilight Bluescan.

(2) http://www.ncbi.nlm.nih.gov/pubmed/8824663

(3) http://justicestudies.sjsu.edu/publications/optimizing-human-semen-stain-detection-using-fluorescence/

(4) http://www.candlepowerforums.com/vb/showthread.php?233945-Cocaine-detection-torch


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