Urine as Forensic Evidence: Identification and Individualisation Techniques

Most people flush it away without a second thought, but urine can quietly decide the outcome of a criminal case. It turns up on bedding in sexual assault investigations, on clothing in cases of confinement or torture, and even in poisoning cases where toxicology needs a sample to work with. Because it looks so ordinary, urine is easy to overlook at a crime scene, yet forensic serologists have built a surprisingly detailed toolkit to spot it, confirm it, and sometimes even trace it back to a specific person. Here’s how that process actually works.

What makes urine useful as evidence 🔗

Urine is a pale yellow, transparent fluid with a sharp, characteristic smell. It is mostly water, making up close to 95 percent of its volume, with the rest made of organic nitrogenous compounds and inorganic salts. Urea is the single largest organic component, alongside smaller amounts of uric acid and creatinine. The yellow colour comes from a pigment called urochrome, and depending on what a person has eaten, taken, or been exposed to, urine can also carry drug metabolites and other chemical traces.

Two physical properties matter a lot in the lab: specific gravity and pH. Specific gravity typically falls between 1.003 and 1.04, while pH can range anywhere from 5 to 8 depending on diet and hydration. Neither figure identifies a stain as urine on its own, but together with the other properties, they give investigators a reasonable starting point before moving to confirmatory chemical tests, as outlined in research on identifying body fluids including urine.

Spotting a urine stain at the scene 🔗

Ultraviolet fluorescence 🔗

Long before any chemical test is run, investigators use ultraviolet light to locate stains that aren’t visible to the naked eye, especially on fabric, mattresses, or carpeting. Under long and short wave UV light, urine stains tend to fluoresce a pale yellow or bluish tone. This isn’t unique to urine, since several other body fluids also fluoresce under UV, but it helps narrow down where to focus closer examination and sampling.

The odour test 🔗

A simpler, low-tech method involves gently heating a suspected stain. Warmth releases the compounds responsible for urine’s distinctive smell, making it easier to detect even in a dried or aged stain. This is only a preliminary check, though, and always needs to be backed up by chemical confirmation before it goes anywhere near a case file.

Confirming it chemically 🔗

Once a stain looks and smells like urine, the next step is proving it through chemistry. Several classic colour and crystal tests target urea, creatinine, and other urinary components.

Urea nitrate crystal test 🔗

This is one of the oldest and most reliable presumptive tests. A small extract of the stain is treated with concentrated nitric acid, which reacts with the urea present to form urea nitrate. Under a microscope, this shows up as distinctive hexagonal, stacked crystals. The shape and arrangement of these crystals is what analysts look for to flag a sample as likely urine, and detailed crystal morphology has been documented in clinical references such as StatPearls’ overview of urinary crystal identification, even though that resource focuses on clinical urinalysis rather than forensic casework.

Gee’s method 🔗

Gee’s method offers another route to detecting urea. It involves extracting the stain with acetone, which helps concentrate the urea present, followed by a reaction that produces visible crystal formation. It’s used as an alternative or supporting test alongside the urea nitrate method, particularly when the stain is old or degraded.

Testing for creatinine 🔗

Creatinine, the second major nitrogenous compound in urine, has its own confirmatory test. When a stain extract is treated with picric acid and sodium hydroxide, a red-orange colour develops if creatinine is present. This colour reaction, sometimes referred to as the Jaffe reaction in clinical chemistry, gives analysts a second, independent marker to support a urine identification alongside the urea-based tests.

The Indian test for indican 🔗

A third confirmatory option is the Indian test, which targets indican, a compound present in urine. The extract is treated with resorcinol reagent followed by cupric bromide, and a positive result produces a distinct red colouration. Running two or three of these tests together, rather than relying on just one, is standard practice because no single colour test is completely specific to urine on its own.

Going beyond identification: whose urine is it 🔗

Confirming that a stain is urine is only the first half of the job. In many cases, investigators also need to know whose urine it is, or at least narrow down the possibilities. This is where things get considerably harder.

Species determination 🔗

Precipitin tests, which use antibody-antigen reactions to tell human samples apart from animal ones, have historically had limited success when applied to urine. Urine simply doesn’t carry the same concentration of proteins that blood does, so classic serological methods struggle. One notable workaround came from Popielski, Brzecka, and Szczecin, who in 1963 described a method that first concentrates the proteins present in the urine sample and then runs them through agar gel double diffusion, a technique closely related to the Ouchterlony test used more broadly for species identification in forensic body fluid analysis. This concentration step made species testing on urine meaningfully more workable than earlier attempts.

ABO blood grouping from urine stains 🔗

Even without nucleated red cells, urine can sometimes reveal a person’s ABO blood group. This works through absorption-elution testing, a technique that detects blood group antigens that are secreted into body fluids by “secretor” individuals. The method has a long history in forensic biology for typing blood, semen, and saliva stains, and the same underlying absorption-elution approach used for ABO typing can be adapted for urine, provided the person is a secretor and the sample hasn’t degraded too much.

The DNA challenge 🔗

DNA profiling is the gold standard in modern forensic identification, but urine is one of the trickier sources to work with. Unlike blood or saliva, urine naturally contains very few nucleated cells, so the DNA yield tends to be low and inconsistent. What DNA is available usually comes from white blood cells and epithelial cells that get shed into the urine, and the amount can vary quite a bit between individuals and even between samples from the same person. Despite these hurdles, researchers have shown that usable genetic profiles can sometimes be recovered, including from dried urine spots tested for individual identification using standard forensic STR kits. This makes urine a viable, if not ideal, backup source of DNA when better samples aren’t available.

Why this matters in real casework 🔗

None of these tests work in isolation. A forensic serologist typically layers physical observation, chemical confirmation, and biological individualisation together, because each method on its own has gaps. UV fluorescence can mislead, colour tests aren’t perfectly specific, and DNA extraction from urine is unreliable. But combined, as described in the broader forensic examination framework for body fluids, they build a case that can hold up to scrutiny in court. This layered approach is exactly why serology training spends so much time on what might otherwise seem like a minor bodily fluid.

What do you think? 🔗

Given how unreliable DNA extraction from urine can be, do you think Indian forensic labs should invest more in refining the older chemical and serological tests, or focus resources on newer DNA recovery techniques instead? And if you were building a crime scene checklist, where would you rank urine stains compared to blood or semen?

References

1. https://www.sciencedirect.com/science/article/abs/pii/S2468170920300357
2. https://www.ncbi.nlm.nih.gov/books/NBK606103/
3. https://ebooks.inflibnet.ac.in/antp07/chapter/429/
4. https://nij.ojp.gov/nij-hosted-online-training-courses/laboratory-orientation-and-testing-body-fluids-and-tissues/testing-body-fluids-tissues/blood-grouping/abo-groups
5. https://pmc.ncbi.nlm.nih.gov/articles/PMC9540579/
6. https://ebooks.inflibnet.ac.in/antp07/chapter/443/