Chemical enhancement of footwear impressions in blood on fabric — Part 3: Amino acid staining

Enhancement of footwear impressions, using ninhydrin or ninhydrin analogues is not considered common practice and such techniques are generally used to target amino acids present in fingermarks where the reaction gives rise to colour and possibly fluorescence. Ninhydrin and two of its analogues were used for the enhancement of footwear impressions in blood on various types, colours and porosities of fabric. Test footwear impressions on fabric were prepared using a specifically built rig to minimise the variability between each impression. Ninhydrin enhancement of footwear impressions in blood on light coloured fabric yielded good enhancement results, however the contrast was weak or non-existent on dark coloured fabrics. Other ninhydrin analogues which have the advantage of fluorescence failed to enhance the impressions in blood on all fabrics. The sequential treatment of impressions in blood on fabric with other blood enhancing reagents (e.g. protein stains and heme reagents) was also investigated.     

Chemical enhancement of footwear impressions in blood on fabric — Part 2: Peroxidase reagents

This study investigates the optimisation of peroxidase based enhancement techniques for footwear impressions made in blood on various fabric surfaces. Four different haem reagents: leuco crystal violet (LCV), leuco malachite green (LMG), fluorescein and luminol were used to enhance the blood contaminated impressions.The enhancement techniques in this study were used successfully to enhance the impressions in blood on light coloured surfaces, however, only fluorescent and/or chemiluminescent techniques allowed visualisation on dark coloured fabrics, denim and leather. Luminol was the only technique to enhance footwear impressions made in blood on all the fabrics investigated in this study.     

A Comparison of Enhancement Techniques for Footwear Impressions on Dark and Patterned Fabrics,

The use of chemical enhancement techniques on porous substrates, such as fabrics, poses several challenges predominantly due to the occurrence of background staining and diffusion as well as visualization difficulties. A range of readily available chemical and lighting techniques were utilized to enhance footwear impressions made in blood, soil, and urine on dark and patterned fabrics. Footwear impressions were all prepared at a set force using a specifically built footwear rig. In most cases, results demonstrated that fluorescent chemical techniques were required for visualization as nonfluorescent techniques provided little or no contrast with the background. Occasionally, this contrast was improved by oblique lighting. Successful results were obtained for the enhancement of footwear impressions in blood; however, the enhancement of footwear impressions in urine and soil on dark and patterned fabrics was much more limited. The results demonstrate that visualization and fluorescent enhancement on porous substrates such as fabrics is possible.     

Chemical enhancement of footwear impressions in urine on fabric

A range of chemical techniques were utilised for the enhancement of footwear impressions deposited on a variety of fabric types of different colours with urine as a contaminant. A semi-automated stamping device was used to deliver test impressions at a set force to minimise the variability between impressions; multiple impressions were produced and enhanced by each reagent to determine the repeatability of the enhancement. Urine samples from different donors were analysed using a spectrofluorophotometer revealing differences between individuals. Results indicated that the enhancement of footwear impressions in urine was possible using amino acid staining techniques whereas protein stains failed to achieve successful enhancement.     

Chemical enhancement of soil based footwear impressions on fabric

This study investigates the enhancement of footwear impressions prepared with soils from different locations on a variety of fabric surfaces with different morphology. Preliminary experiments using seventeen techniques were carried out and the best responding reagents were evaluated further. Results indicated that the soils investigated (a cross-section of soils from Scotland) are more likely to respond to reagents that target iron ions rather than calcium, aluminium or phosphorus ions. Furthermore, the concentration of iron and soil pH did not appear to have an effect on the performance of the enhancement techniques. For the techniques tested, colour enhancement was observed on all light coloured substrates while enhancement on dark coloured fabrics, denim and leatherette was limited due to poor contrast with the background. Of the chemical enhancement reagents tested, 2,2'-dipyridil was a suitable replacement for the more common enhancement technique using potassium thiocyanate. The main advantages are the use of less toxic and flammable solvents and improved clarity and sharpness of the enhanced impression. The surface morphology of the fabrics did not have a significant effect on the enhancement ability of the reagents apart from a slight tendency for diffusion to occur on less porous fabrics such as polyester and nylon/lycra blends.     

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.     

An investigation into the enhancement of fingermarks in blood on fruit and vegetables

A number of studies have reported the successful enhancement of latent fingermarks on fruit and vegetables. A study was set up to identify the most effective technique for the enhancement of fingermarks in blood on various fruit and vegetables. The enhancement techniques targeted different components in blood and consisted of protein stains (e.g. acid black 1), peroxidase reagents (e.g. leuco crystal violet) and amino acid stains (e.g. ninhydrin). Different variables such as the ageing periods of the marks and a diminishing series were employed to assess the suitability and sensitivity of the enhancement techniques.Overall, the use of different protein stains appeared to be the most effective techniques for the enhancement of fingermarks in blood on fruit and vegetables. In addition, the aubergine and cucumber skins appeared to be the most responsive surface to the different chemical techniques during enhancement. On the contrary, little or no enhancement was achieved for fingermarks in blood on the nectarine fruit.     

A preliminary investigation into the use of alginates for the lifting and enhancement of fingermarks in blood

Recent studies have reported the use of alginate in the lifting and subsequent enhancement of footwear marks in blood. A study was set up to assess the use of such a method in the treatment of fingermarks in blood on a variety of porous, non-porous and semi-porous surfaces. Other variables included ageing of the fingermarks in blood and the application of chemicals prior to or post-alginate lifting. All different variations were compared to direct chemical treatment of the substrate. The results demonstrated that alginate is not compatible with certain substrates (e.g. glass and tile). On substrates that were compatible with alginate (e.g. fabric and paper), the enhanced fingermarks on the alginate cast and the enhanced fingermarks on the post-alginate substrates appeared, overall, inferior compared to direct chemical enhancement without the use of alginate. A further variation using water-based protein stains directly mixed with the alginate appeared to provide enhancement directly on the substrate as well as simultaneous lifting and enhancing the fingermarks in blood on the alginate cast.     

The analysis of biological samples from crime scene for a future human DNA profile confrontation. Effects of presumptive test reagents on the ability to obtain STR profiles for human identification

Because of the increase of evidence of blood stains, that have been washed or cleaned in an attempt to mask the analysis of DNA profiles, there is also an increase in the use of presumptive tests on samples sent to laboratories. Some of the presumptive tests, used to identify blood and semen stains, could potentially affect the recovery of high molecular weight DNA from the samples, or extinguish them, especially those already present in small quantities. After the presumptive tests, often these samples are discarded. This study aimed to examine the possibility of obtaining a DNA profile from samples submitted for presumptive testing and cleaned with bleaches with and without chlorine. Two different protocols were conducted: (a) A unique sample of human blood in natura (5 μL), already typed through the DNA techniques with the genetic profile previously known (control), was distributed onto cotton fabrics and dried at room temperature. Four samples of fabric were macerated in saline solution and Coombs serum and then stored for three months (room temperature and freezer −20 °C). (b) Another sample of human blood, type A, in natura, already typed through the techniques of DNA (control) was used. Aliquots of 200 μL were distributed in: cotton, denim and synthetic fabric. The samples were dried at room temperature for 24 h. The blood stains in those fabrics (cotton, denim and synthetic) were then divided into three groups: unwashed, cleaned with chlorine bleach and cleaned with chlorine bleach and soap powder. The samples were again dried at room temperature for 24 h, before the use of luminol. The DNA were extracted with Chelex 100 and amplified with the Identifiler Kit (Applied Biosystems). The blood stains exposed to saline and Coombs serum had DNA profiles consistent with untreated samples (controls). This result shows that the experts should keep and store the samples treated with saline and Coombs serum for future DNA confrontation when necessary. Also discussed in this paper the pattern of blood stains after washing with bleaching solutions, as well as the quantity of DNA obtained from these samples.     

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.     

荧光钙血迹蓝钠试剂增强潜在多介质混合鞋印的应用

目的研究荧光钙血迹蓝钠试剂增强潜在灰尘水渍混合鞋印、灰尘尿渍混合鞋印、灰尘血渍混合鞋印的普适性。方法采用高压微雾雾化法,雾化增强常见的8种地板上的20%w/w灰尘含量的灰尘水渍、灰尘尿渍和灰尘血渍混合鞋印,并与平面扫描提取技术相比较。结果增强后的鞋印花纹清晰,呈亮蓝色,提取效果明显优于自然光下拍照提取,略优于平面扫描提取技术。结论荧光钙血迹蓝钠试剂能够有效增强潜在多介质混合鞋印,是一种便捷高效的潜在混合鞋印增强提取方法。     

The influence of fabric surface characteristics on satellite bloodstain morphology

Bloodstains on fabrics such as clothing, soft furnishings or carpets are often encountered in casework. These stains often have a distinctive morphology that includes satellite stains, thought to be a highly sensitive feature that is a function of surface roughness. This study presents the findings of experimental studies conducted with proxy blood on two fabrics, similar in labeled composition, to assess the influence of fabric type on satellite stain generation. The morphology of proxy blood stains on the two fabric types were found to be statistically distinguishable from one another, with the volume of satellite stains generated being dependent upon the surface roughness of the fabric. These findings provide an initial step that illustrates the viability of providing an empirical evidence base for the interpretation of satellite stains in forensic blood pattern analysis (BPA).     

Chemical enhancement of footwear impressions in blood deposited on fabric — Evaluating the use of alginate casting materials followed by chemical enhancement

Most footwear marks made in blood on a surface such as fabric tend to be enhanced in situ rather than physically recovered using a lifting technique prior to enhancement. This work reports on the use of an alginate material to recover the impressed footwear marks made in blood and deposited on a range of fabric types and colours. The lifted marks were then enhanced using acid black 1 and leuco crystal violet with excellent results.This presents a new method for the lifting and recovery of blood impressions in situ from crime scene followed by subsequent mark enhancement of the lifted impression.     

Human genotyposcopy: the identification of the species, sex and individual by DNA genetic imprints in a case connected with a murder attempt]

Blood stains on a knife were identified by DNA genotyposcopy. The statistical validation method has confirmed that the blood stains on material evidence belonged to the victim, the probability of random coincidence being less than 10(-11). The efficacy of using hypervariable locus-specific DNA probes and the possibility of detecting DNA impressions in blood stains stored for more than 3 months have been demonstrated.     

Systematic study on STR profiling on blood and saliva traces after visualization of fingerprint marks

This paper describes a systematic study of the influence of optical, physical, and chemical methods used for fingerprint enhancement on subsequent DNA analysis of biological stains. Latent fingerprints as well as fingerprints in contact with blood and saliva on different surfaces were treated with dactyloscopic methods. As a general finding, subsequent STR profiling of the blood/saliva traces led to good results after all the enhancement methods included in this study. Concerning blood enhancement procedures, the airbrush technique showed deleterious effects on subsequent STR analysis in some cases. We therefore recommend the implementation of the layer technique, as it brings advantages for fingerprint enhancement as well. It could also be shown that, as can be necessary in practical casework, two enhancement methods can be performed on a single stain without having influence on STR profiling. In terms of methodological variety, this paper reflects a comprehensive study performed on STR profiling after fingerprint enhancement methods, including rare methods and variations of techniques, which can be a useful alternative in certain case scenarios.     

A method for impregnating nylon transfer membranes with leucocrystal violet for enhancing and lifting bloody impressions

The objective of this research project was to demonstrate a quick and easy method for impregnating nylon transfer membranes with leucocrystal violet (LCV) for the purpose of lifting and enhancing impressions made in blood. A stamp that would simulate fine detail found in fingerprints or footwear was used to create impressions on a variety of substrates. Four different LCV formulations were tested to determine the effectiveness of the prepared membranes in lifting and enhancing the impressions. Further investigation involved the feasibility of using the LCV membranes in the field by studying the shelf life and storage of the impregnated membranes and the longevity of the lifted impressions. One of the formations studied demonstrated superior lifting and enhancing capabilities, as well as a prolonged shelf life and a resilience of the lifted impressions, thus proving LCV to be an extremely valuable technique.     

Determination of ABO antigens in fingernails using the APAAP (immunoalkaline phosphatase) technique

Fingernail specimens with adherent nail-bed were taken from autopsy material with blood groups A, AB, B and O. Frozen 4-5-microns sections were submerged and floated carefully during each working step. Portions of fingernails were contaminated with blood and buccal cells, respectively. Furthermore, fingernail fragments of 8 volunteers were embedded in a biocomponent adhesive according to Grieve and Kotowski (Forensic Sci. Soc., 26 29-34) (1986) and cut by the usual microtome technique. APAAP staining is a proper method for demonstrating blood group antigens in fingernails from groove to margin. Frozen sections as well as smallest specimen embedded in a suitable adhesive are applicable for staining procedures. Using freshly prepared artificial stains, blood group constellations of red blood cells and/or buccal cells adherent on the surface of fingernails may be distinguished from the nail matrix.     

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 utility of polyester and cotton as swabbing substrates for the removal of cellular material from surfaces

Various types of cotton and polyester fabrics were tested to ascertain the optimal physical and chemical characteristics of fabrics needed for the removal of cellular material from surfaces. DNA quantitation values obtained on dried saliva stains showed no difference between cotton and polyester across all constructions and solvent conditions. Fabrics used dry and with water yielded higher quantitation values than those used with isopropanol. Quantitation values were also higher for wovens and nonwovens than knits across all solvent conditions. Low thread count fabrics used with water yielded higher quantitation values, but no correlation between thread count and quantitation values was observed with dry fabrics. A low thread count woven fabric, however, outperformed other tested fabrics when swabbing object surfaces in a highly used room. Full DNA profiles from fingerprints on glass surfaces were obtained with low thread count woven and nonwoven fabrics but not with the knit fabric tested.     

Effect of fabric washing on the presumptive identification of bloodstains

The purpose of this study was to investigate the retention of blood stains on twelve different types of fabrics after washing at various drying times. The findings of this study, supported by chi-square analysis, indicate that the retention of bloodstains on washed fabrics depends upon the particular fiber composition of the fabric, the specific blood screening test used, and whether or not a detergent was used in the wash. The results of this research did not reveal a significant effect of the drying time on the retention of bloodstains, as tested during the 48-h limit of this experiment. The author concludes that the forensic serologist should consider the factors investigated in this study before rendering an opinion on the retention of bloodstains on washed garments.