Recent forensic applications of enhanced chromatographic separation methods.

This study reviews the recent applications of enhanced separation methods employed in forensic analysis utilizing gas chromatography, liquid chromatography, and supercritical fluid chromatography published between 2015 to 2020, except papers previously covered in relevant review articles. Applications of enhanced chromatographic separation methods to arson investigations, environmental forensics, sexual assault investigations, drug analysis, and toxicology are discussed. Future directions for enhanced chromatographic separation methods in forensic science are also explored.

Perils of human remains examination in COVID-19 times.

The novel coronavirus, SARS-CoV-2 has surged globally bringing the whole world virtually to a standstill. Due to its highly contagious nature, various guidelines, protocols and preventive strategies have been devised for the safety of healthcare workers during management of the living as well as the dead. However, guidelines and precautions to be followed during the examination of the human skeletal remains are largely lacking. The present communication intends to address the issue of safe handling of human remains during medicolegal investigations in the COVID-19 pandemic times.

Application of forward-looking infrared (FLIR) imaging from an unmanned aerial platform in the search for decomposing remains.

Traditional methods of cadaver detection in outdoor environments include manual ground search, cadaver dogs, and manned aerial reconnaissance during daylight. These methods have limitations; however, a potential low-cost alternative may be to employ thermal imaging equipment mounted on an unmanned aerial vehicle (UAV) to detect heat emitted by insect and bacterial activity on the decomposing remains. No studies have addressed the influence of wrappings on detection of maggot mass thermal signatures nor assessed thermal detection of smaller body fragments. We addressed these knowledge gaps by utilizing a two-phase experimental approach to explore thermal detection of carcasses using UAV-mounted infrared imaging. In Phase 1, pig body fragments were deposited on the surface or shallow buried. In Phase 2, whole pig carcasses were deposited in four conditions: on the surface uncovered, wrapped in plastic or carpet, or buried. Our results demonstrated that observable heat emissions from remains corresponded to peak insect activity during active decay and could be readily detected in uncovered whole carcasses and fragments. Although plastic and carpet wrappings partly impeded detection of insect heat signatures, these materials were clearly detectable themselves because of their contrast to the background ground surface. Thermal signatures of buried partial remains and disturbed gravesoil were also observed; however, the buried whole carcass transitioned to adipocere prior to the decay stage and without any insect colonization or heat signature. These data can inform operational implementation of this technique to complement existing search strategies to offer a robust, low-cost alternative for use where scene characteristics allow.

A comparative analytical study on reagent-fused silica gel plate and wax painted paper-based microfluidic device for serological testing.

The detection and identification of body fluids at a crime scene shed light on the events which might have occurred and the people involved in the crime; the techniques used being lengthy, makes the on-scene detection a tough process to carry out and handle. This study aims to develop an on-spot detection method for serological testing using silica gel encapsulation technique and a modified microfluidic paper-based analytical device (µPADs). Kastle-Meyer reagent was incorporated into the µPAD and Silica Gel plate and was subsequently validated for the detection of blood. This study revealed that the µPAD was a better option. The emerging technique of µPADs allows a cost-effective and simple method of detection for body fluids. The fibrous network of the paper is manipulated to fabricate a guided channel for the fluid flow. The guided channels in the modified µPADs were fabricated by patterning a hydrophobic barrier out of wax. The analyte of interest for this study is blood but the principle can be modified to include other biological fluids.

Blast Suppression Foam, Aqueous Gel Blocks, and their Effect on Subsequent Analysis of Forensic Evidence.

In addition to having blast mitigation properties, aqueous foam concentrate AFC-380 blast suppression foam is designed to capture aerosolized chemical, biological, and radioactive particles during render-safe procedures of explosive devices. Exposure to aqueous environments and surfactants may negatively affect forensic evidence found at the scene, but the effects of AFC-380 foam and aqueous gel on the preservation and subsequent analysis of forensic evidence have not previously been investigated. Sebaceous finger and palm prints and DNA samples on paper, cardboard, tape, and various metal and plastic items, along with hairs, carpet and yarn fibers, and inks and documents, were exposed to AFC-380 foam. Similar mock evidence was also exposed to a superabsorbent gel of the type found in aqueous gel blocks used for shrapnel containment. Exposure to foam or aqueous gel was associated with a dilution effect for recovered DNA samples, but quality of the samples was not substantially affected. In contrast, exposure to AFC-380 foam or gel was detrimental to development of latent finger and palm prints on any substrate. Neither the hair nor the fiber samples were affected by exposure to either the foam or gel. Indented writing on the document samples was detrimentally affected by foam or gel exposure, but not inks and toners. The results from this study indicate that most types of forensic evidence recovered after being exposed to aqueous gel or blast suppression foam can be reliably analyzed, but latent finger and palm prints may be adversely affected.

Evaluation of the use of chemical pads to mimic latent fingermarks for research purposes.

Fingermark detection is in constant evolution, with new techniques being developed and existing ones being continuously optimised. Recently, researchers have begun to express interest in artificial fingermark secretions to overcome the issues arising from the variability of fingermark composition. Some of these artificial secretions have started to appear on the market in the form of pads that can be used to deposit fingermarks with a known and controlled composition. This study aimed at assessing the reliability of three commercially-available pads by comparing the results to those obtained by real fingermarks, using six detection techniques (1,2-indanedione/zinc, ninhydrin, cyanoacrylate followed by rhodamine 6G staining, gold/zinc vacuum metal deposition, and physical developer) on five substrate types (copy and recycled paper, acetate, glass, and glossy paper). The results showed that the artificial fingermarks deposited with these pads reacted in an unreliable way, notably when treated with complex detection techniques such as Physical Developer. Further, the high concentration of some of the target compounds found in the artificial secretion led to an over performance of some detection techniques, which could mislead the operator to overestimating the efficiency of a given method. The resulting artificial fingermarks are considered too dissimilar to real fingermarks to be used as quality control standards and better simulants need to be found for a more efficient and realistic control of the variability.

Cutaneous wound metallic residues pattern analysis with SEM-EDX in a forensic case of overkill.

The term overkill identifies the infliction of massive injuries that greatly exceed those needed to kill the victim both in number and intensity. We present the case of a Peruvian transsexual, who was choked and hit by 11 sharp-force wounds all in the facial area. The scanning electron microscopy with energy dispersion X-ray analysis carried out on the skin of the facial wounds proved crucial in proving that two different point-and-edge weapons were used, following the finding of metallic micro-traces of heterogeneous composition. This discovery and the autopsy highlighted the use of two distinct types of injury (mechanical asphyxia and sharp-force trauma) showed that this murder amounted to overkill.

Capillary zone electrophoresis separation and collection of spermatozoa for the forensic analysis of sexual assault evidence.

The processing of sexual assault kits (SAKs) relies on the genetic analysis of material extracted from swabs collected from the assault victim. A vital step in producing an identifiable DNA profile of the perpetrator is the effective separation of perpetrator (sperm) and victim (epithelial) DNA that have been isolated from the collected evidence. We report the use of capillary zone electrophoresis for the separation of intact sperm from whole and lysed epithelial cells in SAKs. The separated components are deposited into wells of a microtiter plate using a computer-controlled fraction collector, and quantitative PCR is used to verify the collection of sperm cells by targeted amplification of male DNA. We present results from simulated sexual assault samples that have been aged for up to 18 months, as well as vaginal swabs from authentic forensic kits. Components extracted from the vaginal swabs from the SAK comigrated with an aged semen sample at 6.25 ± 0.25 min. Epithelial cells migrated from 10-12 min, producing baseline resolution of the components. Sperm cells were collected in a microtiter plate for downstream analysis.

Thanatochemistry at the crime scene: a microfluidic paper-based device for ammonium analysis in the vitreous humor.

Most of the on-site approaches for inferring of the post-mortem interval are still based on observative data from the direct body inspection, whereas, objective and quantitative analyses, such as potassium in the vitreous humor, are require laboratory instrumentation and skilled personnel. The present paper presents a simple and low cost analytical method suitable for use at the crime scene for inferring the time since death. The method uses a microfluidic paper-based device (µPAD) for the determination of ammonium in the vitreous humor (VH) based on the selective interaction between the ammonium and the Nessler's reagent. The color change was measured in terms of "RGB distance" by using a simple and free smartphone application. The optimized device showed a limit of detection of 0.4 mmol L-1, with between days precision less than 9.3% expressed as relative standard deviation, and accuracy between days from 94.5% to 104.5%. The selectivity of the Nessler's reaction was tested towards the main vitreous humor compounds, and no significant interferences were found. This paper-based analytical device was successfully used for the determination of ammonium ion in VH samples from forensic autopsies. The results obtained with the proposed method, although for a limited number of cases (n = 25), showed a close correlation with the data obtained with an instrumental analysis based on capillary electrophoresis. Moreover, in order to make the evaluation of results as simple as possible, a direct correlation between the color intensity, expressed as RGB distance, and the post-mortem interval was studied and a significant correlation was found (R2 > 0.78). In conclusion, the present preliminary study showes that the proposed device could be an additional tool to the traditional methods for a more accurate, although still presumptive, estimation of the time of death directly at the crime scene.

Differential organic DNA extraction of semen sample contaminated with blood for the identification of a serial sexual offender: A case report.

In many cases of sexual assault, traces of semen are left behind on the victim's body, clothes and the area in which the assault has taken place. The positive identification of semen is instrumental in supporting such cases. There are several methods of forensic examination of semen reported in literature, but the presence of blood complicates the identification of semen stains. This paper presents one such case study where the presence of blood makes DNA profiling more challenging as the PCR amplification becomes complicated, and the absolute differential isolation is the only way to get the clear profile using identifiler kits.

Ratios and distances of pull-up peaks observed in GlobalFiler kit data.

Short tandem repeat (STR) analysis is widely used for forensic examinations with a capillary electrophoresis instrument such as the 3500xL Genetic Analyzer. This instrument adapts multi-locus STR kits to examine up to 27 loci using a 6-fluorescent dye system and corrects the spectral overlap between each dye. However, inaccurate spectral correction can cause pull-up peaks. Here, we examined the pull-up peaks observed in GlobalFiler kit data in terms of their peak height ratios and distances from their parent allele peaks when using the 3500xL and the 3130xl Genetic Analyzers. With the 3500xL, 546 pull-up peaks were observed, and their pull-up ratios averaged 1.03 ±â€¯0.32% (range 0.260-2.80%). Of the 546 pull-up peaks, 534 peaks (97.8%) were within ±1 bp from their parent allele peaks. Overall, the pull-up peaks toward adjacent shorter wavelength channels (e.g., from yellow to green) tended to be observed in the left side (shorter bp) of the corresponding parent allele peaks, and the opposite side tendency was observed for those pull-up peaks toward adjacent longer wavelength channels. These tendencies were also observed in the GlobalFiler data generated with the 3130xl and in the data obtained by injecting a J6 matrix standard with LIZ 500 or 600 v2 size standard into the 3500xL and 3130xl. Inspection of raw data revealed that the shift of pull-up peaks from their parent allele peaks was derived from sigmoid, pull-down, or slightly shifted pull-up shapes. Based on the obtained data, we propose a standard for assessment of questionable pull-up peaks.

Comparison of scanning Kelvin probe with SEM/EPMA techniques for fingermark recovery from metallic surfaces.

Most traditional techniques to recover latent fingermarks from metallic surfaces do not consider the metal surface properties and instead focus on the fingermark chemistry. The scanning Kelvin probe (SKP) technique is a non-contact, non-destructive method, used under ambient conditions, which can be utilised to recover latent prints from metallic surfaces and does not require any enhancement techniques or prevent subsequent forensic analysis. Where a fingermark ridge contacted the metal, the contact potential difference (CPD) contrast between the background surface and the fingermark contact area was 10-50mV. Measurements were performed on the untreated brass, nickel-coated brass and copper metal surfaces and compared to traditional forensic enhancement techniques such as Vacuum Metal Deposition (VMD) using Au-Zn and Au-Ag. Using VMD, the CPD change ranged from 0 to 150mV between the dissimilar metal surfaces affected by the fingermark. In general, SKP worked best without additional enhancement techniques. Scanning Electron Microscope (SEM) scans were used to identify the fingermark contact areas through a sodium, chlorine and oxygen electron probe micro-analyzer (EPMA). The fingermark was observed in the backscattered electron image as the carbon deposits scattered the electrons less than the surrounding metal surface. The fingermark is shown clearly in a Cathodoluminescence scan on the copper sample as it blocks the photon emission at band gap (2.17eV) from the underlying copper oxide (Cu2O) surface. For the first time, SEM, EPMA and Cathodoluminescence techniques were compared to SKP data. Visible and latent fingermarks were tested with latent, eccrinous fingermarks more easily imaged by SKP. Results obtained were very encouraging and suggest that the scanning Kelvin probe technique, which does not need vacuum, could have a place as a first stage analysis tool in serious crime investigation.

[Development of Lung Compression Degree Measurement Software of Pneumothorax and Its Application in Forensic Medicine].

OBJECTIVES: To develop a measurement software of lung compression degree to calculate the lung compression ratio in pneumothorax patients accurately and quickly, and then provide an objective assessment of damage degree in forensic clinical identification. METHODS: A volume calculation software was established according to the working principle of the CT instrument. CT data of 15 pneumothorax patients were selected as research objects. The lung compression ratio of pneumothorax patient was calculated by the lung compression volume calculation software of the CT instrument. Meanwhile, the lung compression ratio was also calculated by the developed volume calculation software. The lung compression ratio and operation time calculated by the two methods were analyzed statistically. Scatter plot graphs were draw based on related data, and the developed volume calculation software was verified. RESULTS: The difference between the lung compression ratios calculated by the two methods was not statistically significant, but showed a linear correlation （P<0.05）. The operation time of the developed volume calculation software was obviously shorter. CONCLUSIONS: The volume calculation software developed in this study can calculate the lung compression degree of pneumothorax more conveniently and rapidly with easy accessibility, which shows an application value in the forensic practice.

The use of ortho-Phenylenediamine and Zar-Pro™ strips for the development of bloodmarks on a dark-coloured, non-porous surface.

The development of fingerprints in blood on a dark-coloured surface can be difficult. Contrast between the bloodmark and the surface can be difficult to achieve using photography and chemical reagents. The use of haem-reagent ortho-Phenylenediamine (OPD) and Zar-Pro™ strips in the enhancement of bloody impressions were investigated. Depletions of bloodmarks were deposited on black ceramic tiles and aged up to 90 days. OPD, Acid Yellow 7, and Hungarian Red were applied to these tiles and the developed fingermarks were compared using a numerical score system. Zar-Pro™ strips were applied to chemically treated and untreated bloodmarks alike. OPD proved to be effective at developing bloodmarks aged up to 90 days, including latent marks. Despite this capability, OPD did not perform as well as Acid Yellow 7 and so the continued use of Acid Yellow 7 is recommended. Zar-Pro™ strips were able to lift bloodmarks aged up to 90days as well as those chemically treated with either: OPD, Hungarian Red, Acid Yellow 7, or Amido Black although ridge detail was poor. Additional investigation into the capabilities of Zar-Pro™ strips for this purpose is recommended.

The evaluation and validation of Phadebas® paper as a presumptive screening tool for saliva on forensic exhibits.

The Phadebas® Forensic Press Test is routinely used for the detection of saliva. However, assessment of the use of Phadebas® paper for this purpose has not been studied extensively. The suitability of Phadebas® paper as a presumptive screening tool for saliva on forensic exhibits, was investigated by analysing the following: (1) sensitivity, (2) specificity, (3) effects of temperature on sensitivity and specificity, (4) detection of saliva in mixed body fluid samples, and (5) influence of substrate porosity. The results of this study demonstrated that Phadebas® paper is more sensitive to α-amylase activity and less specific for saliva than previously reported. The use of an examination temperature of 37°C had no effect on sensitivity, but increased the incidence of cross-reactivity with other forensically relevant body fluid stains. Blood, urine and vaginal secretions can inhibit the detection of α-amylase activity with Phadebas® paper in mixed stains of saliva and body fluid. Substrate porosity is a weak predictor for the time taken for a saliva stain to achieve a strong positive result on Phadebas® paper. Overall, this study demonstrated that the Phadebas® Forensic Press Test has limitations as a presumptive test for the accurate identification of saliva.

Exploitation of microbial forensics and nanotechnology for the monitoring of emerging pathogens.

Emerging infectious diseases remain among the leading causes of global mortality. Traditional laboratory diagnostic approaches designed to detect and track infectious disease agents provide a framework for surveillance of bio threats. However, surveillance and outbreak investigations using such time-consuming approaches for early detection of pathogens remain the major pitfall. Hence, reasonable real-time surveillance systems to anticipate threats to public health and environment are critical for identifying specific aetiologies and preventing the global spread of infectious disease. The current review discusses the growing need for monitoring and surveillance of pathogens with the same zeal and approach as adopted by microbial forensics laboratories, and further strengthening it by integrating with the innovative nanotechnology for rapid detection of microbial pathogens. Such innovative diagnostics platforms will help to track pathogens from high risk areas and environment by pre-emptive approach that will minimize damages. The various scenarios with the examples are discussed where the high risk associated human pathogens in particular were successfully detected using various nanotechnology approaches with potential future prospects in the field of microbial forensics.

Cling film plastic wrap: An innovation for dead body packaging, preservation and transportation by first responders as a replacement for cadaver body bag in large scale disasters.

Cadaver body bags are the conventional method to contain a human body or human remains, which includes the use for storage and transportation of the deceased at any crime scene or disaster scene. During disasters, most often than not, the first responders including the police will be equipped with cadaver body bags to do scene processing of human remains and collection of personal belongings at the disaster site. However, in an unanticipated large scale disasters involving hundreds and thousands of fatalities, cadaver body bags supplies may be scarce. The authors have therefore innovated the cling film plastic wrap as an alternative for the cadaver body bag used at the disaster site. The plastic wrap was tested on six different experimental subjects, i.e. both adult and child mannequins; body parts of the mannequin figure (arm and hand); a human adult subject and an unknown dead body. The strengths of the cling film plastic wrap are discussed in comparison with the cadaver body bag in the aspects of costing, weight, duration of the wrap, water and body fluid resistant properties, visibility and other advantages. An average savings of more than 5000% are noted for both adult body wrap and child body wrap compared to the cadaver body wrap. This simply means that the authors can either wrap 25 adult dead bodies or 80 children dead bodies with the cost of 1 cadaver body bag. The cling film plastic wrap has proven to have significant innovation impact for dead body management particularly by the first responders in large scale disasters. With proper handling of dead bodies, first responders can manage the dead with dignity and respect in an overwhelmed situation to facilitate the humanitarian victim identification process later.

White light reflectance spectroscopy biosensing system for fast quantitative prostate specific antigen determination in forensic samples.

A label-free biosensor based on white light reflectance spectroscopy for the determination of PSA as semen indicator in forensic samples is presented. The sensor is based on a two-step immunoassay which employs the same polyclonal anti-PSA antibody as capture and detection antibody followed by reaction with streptavidin as a signal enhancement step. The whole assay time was set to 10min; 5min reaction of immobilized antibody with the PSA calibrators or the samples, 3min reaction with the biotinylated anti-PSA antibody and 2min reaction with streptavidin. Following this protocol, a detection limit of 0.5ng/mL was achieved and the assay's linear response range extended up to 500ng/mL. Thus, taking into account the quantification limit of 1.0ng/mL and the average PSA concentration in semen (0.2-5.5mg/mL), semen quantities of a few nanoliters could be detected. The accuracy of the sensor developed was demonstrated through recovery (% recovery ranged from 89.6 to 106) and semen dilution experiments. A linear correlation was found for semen dilutions ranging from 5000 to 360,000. The lack of interference by other bodily fluids was confirmed by analysing stains of blood, urine and saliva prior to and after the addition of semen. Finally, the sensor was evaluated by analysing 51 forensic casework samples which were also analysed with a semi-quantitative membrane strip test (Seratec® PSA), through microscopic detection of spermatozoa, and male DNA identification through detection of Y chromosome. The results obtained with the sensor were in excellent agreement with those provided by an immunoradiometric assay kit (PSA-RIACT) and in complete agreement with the findings using the membrane strip assay, spermatozoa and Y chromosome detection. The excellent analytical performance and small size of the instrument make the sensor developed an attractive tool for use in forensic evidence screening for semen detection.

Early Detection of Pressure Injury Using a Forensic Alternate Light Source.

OBJECTIVE: This study aimed to determine if an alternate light source (ALS) can be used to detect tissue trauma before visible manifestations of tissue injury are evident with the naked eye. MATERIALS AND METHODS: Ten participants were recruited and gave consent, and 7 completed the study. Researchers examined and photographed participants' heels in ambient light to establish baseline. A series of photographs using ALS and camera were taken as follows: violet wavelength at 415 nm to 445 nm with yellow lens; blue wavelength at 455 nm to 515 nm with orange lens; and green wavelength at 535 nm to 575 nm with red lens. Participants were examined weekly for 6 consecutive weeks to ascertain skin changes in ambient light and through the ALS. RESULTS: Overt tissue changes were noted when viewed with the ALS and camera compared with visual screens in ambient light. Descriptive statistics were calculated for all wavelengths. Two chi-square tests of independence were run to look for relationships between wavelength and the number of detected injuries (absorption). CONCLUSIONS: Participants presenting with nonblanching erythema in ambient light showed significant tissue absorption under ALS and camera, depicting the actual scope and magnitude of the tissue trauma. Participants with scars, areas of previous injury, and pigmentary changes also showed significant absorption at those sites. These combined findings indicate that ALS can detect tissue trauma and areas at risk not readily visible by the naked eye. This noninvasive tool could help identify patients in the early stages of tissue trauma as well as screen for sites of previous injury that are at risk for subsequent breakdown, saving significant health care dollars and improving outcomes and quality of life.

Current applications of high-resolution mass spectrometry for the analysis of new psychoactive substances: a critical review.

The proliferation of new psychoactive substances (NPS) in recent years has resulted in the development of numerous analytical methods for the detection and identification of known and unknown NPS derivatives. High-resolution mass spectrometry (HRMS) has been identified as the method of choice for broad screening of NPS in a wide range of analytical contexts because of its ability to measure accurate masses using data-independent acquisition (DIA) techniques. Additionally, it has shown promise for non-targeted screening strategies that have been developed in order to detect and identify novel analogues without the need for certified reference materials (CRMs) or comprehensive mass spectral libraries. This paper reviews the applications of HRMS for the analysis of NPS in forensic drug chemistry and analytical toxicology. It provides an overview of the sample preparation procedures in addition to data acquisition, instrumental analysis, and data processing techniques. Furthermore, it gives an overview of the current state of non-targeted screening strategies with discussion on future directions and perspectives of this technique. Graphical Abstract Missing the bullseye - a graphical respresentation of non-targeted screening. Image courtesy of Christian Alonzo.