Discovering Donor Characteristics from Bloodstains with Infrared Spectroscopy

Discovering Donor Characteristics from Bloodstains with Infrared Spectroscopy

From interpreting the incident to pinpointing the perpetrator, the presence of blood at a crime scene can provide clues vital to solving a crime.  Since the advent of DNA profiling in the 1980s, police have been able to use DNA to link suspects to crime scenes, making the detection and collection of biological evidence more important than ever before. However successful DNA profiling relies on a positive match with either a DNA profile from a suspect or one stored in a database. With nothing to compare a profile to, the DNA is of limited use and the trail may quickly run cold.

But what if investigators could gain even more information from a bloodstain at a crime scene? What if it were possible to rapidly figure out whether the donor was male or female, or establish their race? And all of this without shipping samples back to the lab.

New research conducted at the University at Albany in New York has demonstrated that it may be possible to establish some individual donor characteristics in a matter of minutes.

Past research has already demonstrated that the biochemical composition of blood differs between males and females and individuals of different races. But the ability to obtain this information on-site at the crime scene in a matter of minutes could change the way body fluids are processed. In a recent study, Prof. Igor Lednev and his team applied a technique known as attenuated total reflection Fourier transform-infrared (ATR FTIR) spectroscopy to blood analysis, with the aim of establishing whether characteristics such as sex and race can be determined from bloodstains.

FTIR is an analytical technique capable of providing information about a material’s chemical information. In brief, the device directs infrared radiation towards the sample. Some of this radiation is absorbed by the material, and some passes through. The sample’s absorbance of this light at different wavelengths is measured and used to determine the material’s chemical information. After analysis a spectrum is produced, which acts as a kind of molecular ‘fingerprint’ of the sample. The different features in the spectrum relate to the different chemical components in the sample.

Infrared spectra were produced by analysing the blood of 30 donors (a mixture of male and females of Caucasian, African American and Hispanic racial origin). From this, researchers could observe any differences occurring between blood from male and females, and blood from members of different races. Using this data, the researchers built a model capable of classifying samples based on their chemical profile. By taking the chemical profile of an unknown bloodstain and comparing it with a model containing bloodstains from numerous different groups, the model can predict the likely classification (i.e. whether the donor was male or female and which racial group they belong to). In this study, it correctly classified bloodstains around 90% of the time.

Using infrared-based techniques has a number of advantages over other methods of analysis. As the technique simply necessitates the direction of light towards the bloodstain, the technique is non-destructive. Inevitably this is perfect for criminal investigations – destroying the evidence is never ideal. IR spectroscopy is also amenable to portability, lending itself well to on-the-go analysis at crime scenes and so potentially saving a lot of time by avoiding sending unnecessary samples back to the lab for analysis.

Although only a pilot study, this research has demonstrated the possibility of establishing donor characteristics through the rapid and non-destructive analysis of bloodstains. The ability to determine features such as sex and race would enable police to significantly narrow down the search for suspects or victims, ultimately preserving valuable time and money. Furthermore, the ability of FTIR to non-destructively analyse evidence on-site renders it an ideal tool for forensic analysis. Inevitably a great deal more research will be necessary, and if the technique ever becomes operational, it would be years before such technology and methods were suitable for deployment to crime scenes and use as evidence in criminal trials.


Mistek et al. Phenotype profiling for forensic purposes: nondestructive potentially on scene attenuated total reflection Fourier transform-infrared (ATR FT-IR) spectroscopy of bloodstains. Forensic Chemistry. 2019, In Press.


Caught Red-Handed: MALDI Mass Spectrometry & Bloodied Fingerprints

Caught Red-Handed: MALDI Mass Spectrometry & Bloodied Fingerprints

Most previous methods of establishing whether a fingermark at a crime scene contain blood are purely presumptive. The suspected fingermark, whether it be a print merely contaminated with traces of blood or an entire mark left in blood, will be subjected to tests which will aim to confirm or refute the presence of blood. However most existing presumptive tests suggest that it is a possibility the fingermark in question contains blood… but that it equally could be another similar substance that happens to produce a positive response with the test used. Thus is the limitation of any presumptive test – they can only give a possibility, not a definitive answer. Obviously not ideal during a forensic investigation.

Suspected bloodstains can be subjected to a wide range of tests to ascertain their composition. Blood may be visualised using alternative light sources, but this is a far cry from confirming its composition and in some cases (such as with the use of short-wave ultraviolet light) can even be destructive to DNA, thus obviously not ideal for the forensic examination of a blood sample. Spectroscopic techniques such as Raman spectroscopy have proved successful in potentially distinguishing blood from other biological fluids, though this has not been widely applied, particularly to blood in fingermarks. Chemical enhancement techniques have also been developed in the past, such as those that react with amino acids or haem-reactive compounds present to produce a colouring or fluorescence to enhance the blood. As successful as these methods may have been in the past, they are still only presumptive and cannot claim with any kind of near-certainty that any positive reaction produced is the result of blood and furthermore whether that blood is of human origin.

As a result of this, more confirmatory tests are needed.

More affirmative procedures do exist and are currently being developed. A particularly interesting method of detecting blood in fingermarks is using a technique known as MALDI MS. That is, Matrix-Assisted Laser Desorption Ionisation Mass Spectrometry. This relatively new analytical technique (relative to the history of mass spectrometry anyway) is most commonly applied to determining the mass of peptides, proteins and polymers, so is ideal for focusing on certain components of blood.

For those unfamiliar with mass spectrometry, in its simplest form it is a technique which is used to determine the identity of sample components based on their mass-to-charge ratio and, in some cases, how the molecule fragments when ionized. MALDI is something of a variation of this technique. In this technique, the sample to be analysed is mixed with a particular matrix material and applied to a plate. A laser irradiates the sample and matrix, causing ablation and desorption, after which the sample is ionized and then accelerated and detected using mass spectrometry.

Researchers have applied MALDI MS to detecting the presence of blood by specifically focusing on the detection of haem and haemoglobin molecules based on their mass-to-charge ratios. These molecules are vital components of blood, with haemoglobin being the protein responsible for oxygen transportation and haem being a compound embedded into haemoglobin which provides the iron essential for oxygen binding. By subjecting known and suspected blood stains and bloodied fingermarks to this technique, haemoglobin chains could be detected even in traces of blood invisible to the naked eye. Initial research into this technique studied human, equine and bovine haemoglobin, establishing that it is possible to determine whether or not haemoglobin was from a human source using mass spectrometry at a high mass range. Both fresh and aged blood samples could be successfully analysed, making the application potentially beneficial to samples from various points in time. Furthermore, the technique has proven to be compatible with other methods often used by investigators when attempting to enhance fingermarks at incident scenes, meaning the new method is not likely to interfere with existing procedures.

A typical haemoglobin molecule.

A typical haemoglobin molecule.

This fascinating application of matrix-assisted laser desorption ionisation mass spectrometry offers a whole new world of possibilities in blood detection in forensic science. Although at present such instrumentation is far from being the norm in the forensic scientist’s arsenal, the applications of advanced mass spectrometry techniques to answering some of the simpler yet vital questions during a criminal investigation make for a captivating read.


Bradshaw, R et al. Direct detection of bloos in fingermarks by MALDI MS profiling and imaging. Science and Justice. 45 (2014), pp. 110-117.

King’s College London. An Introduction to Mass Spectrometry Based Proteomic. [online][Accessed 16 Feb 2015] Available: