Identification and age estimation of blood stains on colored backgrounds by near infrared spectroscopy

Non-destructive identification and subsequent age estimation of blood stains are significant steps in forensic casework. The latter can provide important information on the temporal aspects of a crime. As previously shown, visible spectroscopy of blood stains on white backgrounds can successfully be used for their identification and age estimation. The use of this technique however, is hampered by dark backgrounds. In the present study the feasibility to use near infrared (NIR) spectroscopy was evaluated for blood stain identification and age estimation on dark backgrounds. Using NIR reflectance spectroscopy, blood stains were distinguished from other substances with 100% sensitivity and 100% specificity. In addition, Partial Least Squares Regression analysis was applied to estimate the age of blood stains on colored backgrounds. The age of blood stains up to 1 month old was estimated successfully with a root mean squared error of prediction of 8.9%. These findings are an important step toward the practical implementation of blood stain identification and age estimation in forensic casework, where a large variety of backgrounds can be encountered.     

The application of visible wavelength reflectance hyperspectral imaging for the detection and identification of blood stains

Current methods of detection and identification of blood stains rely largely on visual examination followed by presumptive tests such as Kastle–Meyer, Leuco-malachite green or luminol. Although these tests are useful, they can produce false positives and can also have a negative impact on subsequent DNA tests. A novel application of visible wavelength reflectance hyperspectral imaging has been used for the detection and positive identification of blood stains in a non contact and non destructive manner on a range of coloured substrates. The identification of blood staining was based on the unique visible absorption spectrum of haemoglobin between 400 and 500 nm. Images illustrating successful discrimination of blood stains from nine red substances are included. It has also been possible to distinguish between blood and approximately 40 other reddish stains. The technique was also successfully used to detect latent blood stains deposited on white filter paper at dilutions of up to 1 in 512 folds and on red tissue at dilutions of up to 1 in 32 folds. Finally, in a blind trial, the method successfully detected and identified a total of 9 blood stains on a red T-shirt.     

Visualization of Latent Blood Stains Using Visible Reflectance Hyperspectral Imaging and Chemometrics

The detection of latent traces is an important aspect of crime scene investigation. Blood stains on black backgrounds can be visualized using chemiluminescence, which is invasive and requires a darkened room, or near-infrared photography, for which investigators need to change filters manually to optimize contrast. We demonstrated the performance of visible reflectance hyperspectral imaging (400–720 nm) for this purpose. Several processing methods were evaluated: single wavelength bands, ratio images, principal component analysis (PCA), and “SIMPLe-to-use Interactive Self-modeling Mixture Analysis” (SIMPLISMA). Using these methods, we were able to enhance the contrast between blood stains and 12 different fabrics. On black cotton, blood dilutions were visible with a minimal concentration of 25% of whole blood. The hyperspectral camera system used in this study is portable and wireless, which makes it suitable for crime scene use. The described technique is noncontact and nondestructive, so all traces are preserved for further analysis.     

Forensic Identification of Blood in the Presence of Contaminations Using Raman Microspectroscopy Coupled with Advanced Statistics: Effect of Sand,Dust, and Soil

Body fluid traces recovered at crime scenes are among the most common and important types of forensic evidence. However, the ability to characterize a biological stain at a crime scene nondestructively has not yet been demonstrated. Here, we expand the Raman spectroscopic approach for the identification of dry traces of pure body fluids to address the problem of heterogeneous contamination, which can impair the performance of conventional methods. The concept of multidimensional Raman signatures was utilized for the identification of blood in dry traces contaminated with sand, dust, and soil. Multiple Raman spectra were acquired from the samples via automatic scanning, and the contribution of blood was evaluated through the fitting quality using spectroscopic signature components. The spatial mapping technique allowed for detection of “hot spots” dominated by blood contribution. The proposed method has great potential for blood identification in highly contaminated samples.     

Highly Sensitive Detection of Blood by Surface Enhanced Raman Scattering,,

Raman spectroscopy for forensic body fluid analysis has received some attention due to the nondestructive nature and potential application for identification at the crime scene; however, its usage has been limited by low detection sensitivity. Surface enhanced Raman scattering (SERS) was evaluated for blood identification for forensic applications. Specifically, a SERS‐active substrate was fabricated, composed of nickel nanotips coated with Ag nanoparticles. Compared with a conventional substrate, the SERS substrate enhanced Raman scattering by more than two orders of magnitude and allowed blood to be identified to a dilution of 1:100,000. Blood was also successfully detected by swabbing the SERS substrate directly on mock evidence. Most importantly, Raman spectra obtained by swabbing the SERS substrate on blood stains were free of luminescence even when blood was deposited on luminescent fabrics. The nondestructive character, simplicity of sample preparation, and high sensitivity make SERS a prime candidate for field and laboratory‐based blood identification.     

The use of Polilight in the detection of seminal fluid, saliva, and bloodstains and comparison with conventional chemical-based screening tests

Biological stains can be difficult to detect at crime scenes or on items recovered from crime scenes. The use of a versatile light source may assist in their detection. The ability of Polilight to locate potential semen, saliva, and blood stains on a range of substrates and at different dilutions was tested. We also tested the use of Polilight in comparison with conventional chemical-based presumptive screening tests such as acid phosphatase (AP), Phadebas, and luminol, often used in casework for detecting potential semen, saliva, and blood stains, respectively. The Polilight was able to locate stains that were not apparent to the naked eye. The color of the material on which a stain is deposited can have an effect on the detectibility of the stain. The Polilight was found to be comparable with the AP and Phadebas tests in terms of its sensitivity. In a comparative study between the AP test and Polilight on 40 casework exhibits, one false-negative result was observed when using the Polilight. On a series of mock casework exhibits it was determined that the Polilight can be used successfully to locate saliva stains for DNA analysis. The sensitivity of luminol for detecting potential bloodstains was greater than that of Polilight; however the Polilight has particular application in instances where a bloodstain may have been concealed with paint. Overall, the Polilight is a relatively safe, simple, noninvasive, and nondestructive technique suitable for use in forensic casework.     

Volume Determination of Fresh and Dried Bloodstains by Means of Optical Coherence Tomography

The volume of bloodstains found on crime scenes may help forensic investigators reconstruct the location and kinematics of bloodletting events, as stain size, volume, and impact velocity are related. Optical coherence tomography was used as a method to determine the volume and volume ratio of dried and fresh bloodstains on both glass and irregular surfaces or deposited with an impact velocity. The volume of blood drops deposited on smooth glass surfaces was measured within a deviation of 2%. This deviation increased for droplets on irregular surfaces or deposited with an impact velocity. The volume ratio of dried and fresh bloodstains was equal to 19–28% depending on the individual donor and on the use of an anticoagulant. Optical coherence tomography is a good method to determine the volume of fresh and dried bloodstains in laboratory conditions and allows accurate determination of the dry/fresh ratio.     

Raman spectroscopy offers great potential for the nondestructive confirmatory identification of body fluids

Raman spectroscopy was used to compare body fluids commonly found at crime scenes in a nondestructive manner. The dry traces of semen, vaginal fluid, sweat, saliva, and blood were analyzed using confocal Raman microscopy with a 785-nm excitation. The results show that the five fluids can be differentiated from one another by visual comparison of their Raman spectra, and that the laser radiation does not damage the sample. The Raman signature of each body fluid is specific and correlates with the known composition of the fluid. Dry traces of human and canine semen exhibited distinctly different Raman signatures. Overall, this preliminary study demonstrates the great potential of Raman spectroscopy for nondestructive, confirmatory identification of body fluids for forensic purposes.     

Genetic identification of degraded DNA samples buried in different types of soil

Biological samples buried in different types of soil are often found in crime scenes. These samples are usually highly degraded which difficult their analysis. Several factors contribute to the degradation of biological material including temperature variation, humidity, UV light and especially the presence of microorganisms.Blood was collected from three non-related male donors and blood stains were made in fabrics such as jeans, cotton and lycra. Blood stains were dried at room temperature and buried in three different types of soil (sand, marsh and clay), to promote its natural degradation.The buried samples suffered a high degradation over time which difficult their genetic identification. The marshy soil proved to be the most destructive one, leading to rapid degradation of the different analyzed fabrics, which disabled the obtainment of the genetic profiles.     

A comprehensive ATR-FTIR spectroscopic analysis for the identification and differentiation of lip balms

Lip balm may be encountered as physical evidence in cases involving sexual assaults, homicides, and kidnappings. Lip balm can be used as corroborative evidence by providing a potential link between the victim, accused, and the crime scene. For lip balms to be used as evidence, it is important to understand the diversity and their aging process under different conditions. Therefore, in this study, ATR-FTIR spectroscopy in conjunction with chemometric tools such as principal component analysis (PCA) and linear discriminant analysis (LDA) has been used for the objective identification and differentiation of 20 brands of lip balms. Moreover, lip balms on different substrates and wearing effects over time were also investigated. The results show that the PCA-LDA training accuracy was 92.5%, whereas the validation accuracy comes out to be 83.33%. A blind study using pristine samples was also performed which resulted in 80% PCA-LDA accuracy. PCA-LDA prediction of samples on various substrates showed a higher chemometric prediction accuracy for nonporous substrates (glass, plastic, and steel), than for porous substrates (cotton cloth, cotton swab stick, dry tissue paper, and white paper) for samples kept in room temperature and under sunlight for 15 days. The substrate study showed that the samples from various substrates could effectively generate respective spectra which can help in brand-level identification even after several days. The present method demonstrates a potential for lip balm samples to be used in forensic casework applications.     

Shoeprint image retrieval and crime scene shoeprint image linking by using convolutional neural network and normalized cross correlation

A shoeprint image retrieval process aims to identify and match images of shoeprints found at crime scenes with shoeprint images from a known reference database. It is a challenging problem in the forensic discipline of footwear analysis because a shoeprint found at the crime scene is often imperfect. Recovered shoeprints may be partial, distorted, left on surfaces that do not mark easily, or perhaps come from shoes that do not transfer marks easily. In this study, we present a shoeprint retrieval method by using a convolutional neural network (CNN) and normalized cross-correlation (NCC). A pre-trained CNN was used to extract features from the pre-processed shoeprint images. We then employed NCC to compute a similarity score based on the extracted image features. We achieved a retrieval accuracy of 82% in our experiments, where a “successful” retrieval means that the ground truth image was returned in the top 1% of returned images. We also extend our shoeprint retrieval method to the problem of linking shoeprints recovered from crime scenes. This new method can provide a linkage between two crime scenes if the two recovered shoeprints originated from the same shoe. This new method achieved a retrieval accuracy of 88.99% in the top 20% of returned images.     

Evaluation of Direct PCR Amplification Using Various Swabs and Washing Reagents

DNA profiles were generated via direct amplification from blood and saliva samples deposited on various types of swab substrates. Each of the six non-FTA substrates used in this research was punched with a Harris 1.2 mm puncher. After 0.1 μL of blood or 0.5 μL saliva, samples were deposited on each of these punches, samples were pretreated with one of four buffers and washing reagents. Amplification was performed using direct and nondirect autosomal and Y-STR kits. Autosomal and Y-STR profiles were successfully generated from most of these substrates when pretreated with buffer or washing reagents. Concordant profiles were obtained within and between the six substrates, the six amplification kits, and all four reagents. The direct amplification of substrates which do not contain lysing agent would be beneficial to the forensic community as the procedure can be used on evidence samples commonly found at crime scenes.     

Impact Spatter Bloodstain Patterns on Textiles

There are few reports of studies of impact spatter on textiles even though bloodstained textiles are found at many violent scenes. Impact spatter was deposited at 90° impact angle onto three knit fabrics of different yarn sizes and on paper. The resulting stain areas and number of stains were measured using ImageJ and compared with stains on paper using one‐factor ANOVA. The number of stains observed and their areas on the knit fabrics decreased as the yarn size increased. It was also found that blood that deposited on the fabric wicked only in the direction of the fibers at that location within the fabric which led to distorted stain shapes. Fewer observed impact spatter stains were found on cotton jersey knits for fabrics made with larger yarns than on paper. As the yarn size became smaller, the number of stains became the same as on paper.     

Rapid and nondestructive analysis of lipstick on different substrates using ATR-FTIR spectroscopy and chemometrics

Lipstick can be an important piece of evidence in crimes like murders, rapes, and suicides. Due to its prevalence, it can be an important corroborative evidence in crime reconstruction. The analysis of such evidence can provide an evidentiary link between the suspect, the victim, object, or the crime scene. We report the use of nondestructive ATR-FTIR spectroscopy combined with chemometrics for the classification of 10 brands of lipsticks with nine samples each. Chemometric method of partial least square-discriminant analysis (PLS-DA) has been employed to interpret the data and classify the samples into their respective classes. The PLS-DA model provides an AUC figure above 0.99 in all brands except one; for which it is slightly less at 0.94. We have also tested the traces of these lipstick samples on different substrates treating them as unknowns in the already trained PLS-DA model. 100% of the samples on nine substrates viz. a cotton, nylon, plastic, dry tissue, denim (blue jeans), wet tissue, nitrile gloves, white paper, and polyester were correctly attributed to their source brand. In conclusion, the results suggest that ATR-FTIR combined with the chemometrics is a rapid, nondestructive, and accurate method for the discrimination and source attribution of lipstick. This study has potential for use in actual forensic casework conditions.     

The ability of the blowflies Calliphora vomitoria (Linnaeus), Calliphora vicina (Rob-Desvoidy) and Lucilia sericata (Meigen) (Diptera: Calliphoridae) and the muscid flies Muscina stabulans (Fallén) and Muscina prolapsa (Harris) (Diptera: Muscidae) to colonise buried remains

A novel method for the non-destructive age determination of a blood stain is described. It is based on the measurement of the visible reflectance spectrum of the haemoglobin component using a microspectrophotometer (MSP), spectral pre-processing and the application of supervised statistical classification techniques. The reflectance spectra of sample equine blood stains deposited on a glazed white tile were recorded between 1 and 37 days, using an MSP at wavelengths between 442 nm and 585 nm, under controlled conditions. The determination of age was based on the progressive change of the spectra with the aging of the blood stain. These spectra were pre-processed to reduce the effects of baseline variations and sample scattering. Two feature selection methods based on calculation of Fisher's weights and Fourier transform (FT) of spectra were used to create inputs into a statistical model based on linear discriminant analysis (LDA). This was used to predict the age of the blood stain and tested by using the leave-one-out cross validation method. When the same blood stain was used to create the training and test datasets an excellent correct classification rate (CCR) of 91.5% was obtained for 20 input frequencies, improving to 99.2% for 66 input frequencies. A more realistic scenario where separate blood stains were used for the training and test datasets led to poorer successful classification due to problems with the choice of substrate but nevertheless up to 19 days a CCR of 54.7% with an average error of 0.71 days was obtained.     

Defining event reconstruction of digital crime scenes

Event reconstruction plays a critical role in solving physical crimes by explaining why a piece of physical evidence has certain characteristics. With digital crimes, the current focus has been on the recognition and identification of digital evidence using an object's characteristics, but not on the identification of the events that caused the characteristics. This paper examines digital event reconstruction and proposes a process model and procedure that can be used for a digital crime scene. The model has been designed so that it can apply to physical crime scenes, can support the unique aspects of a digital crime scene, and can be implemented in software to automate part of the process. We also examine the differences between physical event reconstruction and digital event reconstruction.     

Measuring the impact of forensic science in detecting burglary and autocrime offences

As UK investment in forensic science has increased, the government has taken a fresh interest in how far this has led to dividends in terms of the detection of crime and its reduction. The Home Office funded 'Pathfinder Project' sought to monitor and document the complex relationships between the collection and use of forensic material (looking at a range of forensic science techniques) and its impact on crime detection. The project specifically targeted the 'volume' crimes of burglary and vehicle crime. Detailed data was gathered on all stages of the process between the collection and use of forensic material and crime detection. The model falls into two conceptual phases--scene attendance to suspect identification and identification to detection. From the analysis it was found that approximately one third of burglary and autocrime scenes are visited by SOCOs. While scientific identifications are only made in a minority of burglary and autocrime offences overall, it belies their importance. About one in ten of burglary and autocrime cases are cleared up by the police and it is estimated that fingerprints and SGMPlus were a contributory factor in achieving one third of these clear ups.     

Human semen stain analysis in casework sample by HRM-qPCR

Determining the biological origin of body fluid evidence from crime scenes is extremely important for criminal investigation, especially in sexual assault cases. In Brazil, sexual crimes still present low-resolution rates, where approximately 8% of cases are judged. The determination of the presence of semen in samples from crime scenes as a test prior to DNA analysis is a mandatory requirement in forensic analysis and can help to better understand the dynamics of the event. This report aims to present the methodological strategy used in a criminal case of a home invasion where a t-shirt containing visible stains similar to human semen was found at the site. Convencional tests to detect the presence of PSA and sperm cells were performed on the fabric cutouts which showed negative results. We then processed the fabric samples for genetic analysis after two-years-storage, where were performed automated method for the genetic material isolation (QIAcube, Qiagen). The Real-time PCR analysis were carried out using the SOLIScript 1-step SolisGreen (SolisBiodyne) kit, with specific primers to the TGM4 (Transglutaminase 4) gene, in the Rotor Gene Q-5Plex HRM equipment (Qiagen). The results obtained for the melt curve indicated the presence of human semen in the analyzed samples. After the HRM-qPCR assay we also analyzed the a-STR and Y-STR markers. All the results were useful for the criminal process, which led to the identification of the author of the crime.