Two-tailed delimitation of the time of death by immunohistochemical detection of somatostatin and GFAP

Delimitating the time of death becomes increasingly difficult and inaccurate, the further back it lays in time. To contribute to available techniques, pancreas and brain tissue from 500 corpses, whose time of death lay within 1 day and 23 days ±1 day, were immunohistochemically stained with anti-somatostatin and anti-glial fibrillary acidic protein (GFAP) antibodies. Somatostatin and GFAP are recognized as antigen by the employed antibodies. This does not occur when proteolytic processes after death disturb the tertiary structure, altering its antigen properties. In our cases, stainability of somatostatin in the pancreas was always given within 2 days after death. Hence, given a negative immunoreaction, death can be assumed to have occurred a minimum of 3 days before formaldehyde fixation, which stops the proteolysis. A negative immunoreaction occurred consistently after 11 days, indicating that if a positive reaction is obtained in a specimen, the death of the respective person must have occurred amaximum of 10 days before fixation. GFAP was always stainable in the frontal cortex within 3 days after death, which means that in the case of a negative immunoreaction, death can be assumed to have occurred a minimum of 4 days before fixation. A negative immunoreaction occurred consistently after 14 days, indicating that if a positive reaction is obtained in a specimen the death of the respective person must have occurred a maximum of 13 days before fixation. The presented methods provide further improvements of the possibilities of delimitating the time of death, whereby the combination of both methods allows a two-tailed delimitation.     

Photometric measurement of color changes in livor mortis as a function of pressure and time

Summary Until now, blanching of livor mortis in estimating time of death has generally been assessed based on subjective impressions, i.e. on whether blanching is visible after the application of pressure. We have developed a measuring system that uses digital processing to objectify the relationship between the pressure applied and blanching of postmortem lividity. The pressure is electronically registered by a strain gauge. At predefined levels (10, 20, 30, up to 100N) the software triggers a color measurement by a commerically available tristimulus colorimeter. All parameters are measured in a single procedure and routed to the computer through a data interface. The pressure-induced color changes in the livor mortis are evaluated according to the L*, a*, b* system (CIE-LAB according to DIN 5033, DIN 6174), which closely approximates the physiology of sight. An additional color spacing formula (AE) allows analysis of color changes irrespective of the basic skin tone. Initial measurements on cadavers showed that application of increasing pressure produced regular courses of color changes in livor mortis.     

Determination of the time of death by fungal growth

Summary A woman was found in her bedroom, which had been kept at a constant temperature of 12°C, several weeks after she had been murdered. The establishment of the time of death was attempted by examination of fungal development on two parts of her body. Agar slopes were inoculated with the fungal growth present on an eyelid and on the inguinal skin. After 1 day at 20°C, the cultures were incubated at 11–12°C. The cultures were then observed daily until growth was comparable to that on the tissues that had been frozen. By these means it could be established that the victim died at least 18 days before her body was discovered. This, in fact, turned out to be the case. Hyphomycetes growth on a corpse may be helpful in determining the time of death when the event happened 10–20 days earlier, provided that the average surrounding temperature is known.     

Rectal temperature-based death time estimation in infants

In determining the time of death in infants based on rectal temperature, the same methods used in adults are generally used. However, whether the methods for adults are suitable for infants is unclear. In this study, we examined the following 3 methods in 20 infant death cases: computer simulation of rectal temperature based on the infinite cylinder model (Ohno’s method), computer-based double exponential approximation based on Marshall and Hoare’s double exponential model with Henssge’s parameter determination (Henssge’s method), and computer-based collinear approximation based on extrapolation of the rectal temperature curve (collinear approximation). The interval between the last time the infant was seen alive and the time that he/she was found dead was defined as the death time interval and compared with the estimated time of death. In Ohno’s method, 7 cases were within the death time interval, and the average deviation in the other 12 cases was approximately 80 min. The results of both Henssge’s method and collinear approximation were apparently inferior to the results of Ohno’s method. The corrective factor was set within the range of 0.7–1.3 in Henssge’s method, and a modified program was newly developed to make it possible to change the corrective factors. Modification A, in which the upper limit of the corrective factor range was set as the maximum value in each body weight, produced the best results: 8 cases were within the death time interval, and the average deviation in the other 12 cases was approximately 80 min. There was a possibility that the influence of thermal isolation on the actual infants was stronger than that previously shown by Henssge. We conclude that Ohno’s method and Modification A are useful for death time estimation in infants. However, it is important to accept the estimated time of death with certain latitude considering other circumstances.     

Morphology of sweat glands in determining time of death

This study demonstrates post-mortem autolytic alterations in the skin at cellular and subcellular levels and identifies parameters which may assist in determining the time of death in the first few hours post-mortem. Serial skin samples from the ventral surface of the arm were taken at intervals of 3, 6, 9 and 12 h after death in 29 subjects of various ages, with no signs of skin disease; causes of death were various. Three types of tests were performed: cytochemical (hematoxylin-eosin and alcian-PAS), immunohistochemical (S-100, CEA, Cytokeratin, ASM) and ultrastructural (electron microscopy). Electron microscopy proved useful for identifying transformations which were found to be specific for each chronological step considered: reduction of intracellular glycogen in clear cells and reduction of secretory granules in dark cells are typcial signs of the first stage (3 h) after death; mitochondrial dilatation and rarefaction of cristae in clear and dark cells are typical of the second stage (6 h); rarefaction of microvilli in dark and clear cells is a sign of the last stage (12 h). Cytochemistry and immunohistochemistry supply useful information — not for all the chronological stage considered here, but for individual phases (3 h for hematoxylin-eosin and 6 h for alcian-PAS). However, it is particularly important to use the results from all such techniques simultaneously, so that the question of the exact time of death within the first 12 h post-mortem may be more accurately answered.     

Estimation of time since death from CSF electrolyte concentration in Bhopal region of central India

Estimation of time since death from sodium and potassium ion concentration levels in CSF (cerebrospinal fluid) was carried out in 100 medico legal autopsies with known time of death in the department of Forensic medicine, Gandhi Medical College in Bhopal region of Central India. CSF was aspirated from lateral ventricles after opening the skull and dura, and concentration of these ions were estimated by flame photometry. Results revealed a significant correlation of sodium and potassium ions in CSF up to 25 h of time since death, with average per hour rise of 1.21 meq/h for potassium and fall of 1.115 meq/h for sodium ions. A useful relationship between sodium potassium ion ratio and PMI (post-mortem interval) was also elicited. The study concludes that changes in CSF electrolyte is a significant parameter to estimate time since death.     

猪椎体火器伤早期死亡原因的实验研究

目的　建立家猪L2 椎体贯通火器伤实验模型,通过监测生命体征等指标变化及弹道损伤情况,分析高速椎体火器伤导致动物早期死亡的原因。方法　18只家猪随机分为2组:低速枪伤组(n =10 ) ;高速枪伤组(n =8)。全麻状态下建立L2 椎体侧方贯通伤模型,分别于伤前、伤后0 .5、1、2和3小时行血气分析、平均动脉压及脉率监测和损伤部位弹道CT扫描。结果　18只家猪中13只成功建立了L2 椎体火器伤模型,其中低速枪伤组6只、高速枪伤组7只。不同时相点2组动物平均动脉压进行性下降、心率进行性加快,但高速枪伤组较低速枪伤组变化幅度较大(P <0 .0 1) ;2组动物血pH、细胞外碱剩余(BE ecf)、血液碱剩余(BE B)、标准碳酸根浓度(SB)、实际碳酸氢根(AB)、氧分压(PaO2 )、二氧化碳分压(PaCO2 )和氧饱和度(SaO2 )呈进行性下降,但高速枪伤组出现的时间早、下降的幅度大(P <0 .0 5或0 .0 1) ;低速枪伤组动物3.5小时死亡率为16 .6 7%、高速枪伤组为85 .71% ,平均死亡时间分别为(7.1±1.1)小时和(3.5±0 .5 )小时,2组比较有显著性差异(P <0 .0 5和0 .0 1)。结论　枪弹的速度是决定损伤程度的重要决定因素之一,高速火器伤动物早期死亡的主要原因是弹道周围广泛创伤所致的失血性休克。     

Postmortem activity of lactate and malate dehydrogenase in human liver in relation to time after death

Summary Changes in the activity of lactate (EC 1.1.1.27, LDH) and malate (EC 1.1.1.37, MDH) dehydrogenases were measured in tissue extracts of human liver kept at 5 different temperatures until 35 days after death. The investigated activities decreased in proportion to time of storage which enabled an estimation of time after death by statistical analysis of the data.     

Application of a simple enzymatic digestion method for diatom detection in the diagnosis of drowning in putrified corpses by diatom analysis

The reliability and applicability of quantitative and qualitative diatom analysis by an enzymatic digestion method in the diagnosis of drowning of putrified bodies has been evaluated. The authors report the analysis of water and organ samples of 12 immersion cases using light microscopy. This study included control organ samples from the bodies of persons who died from causes other than drowning. Organ samples were treated by both chemical and enzymatic methods, the first one using concentrated nitric acid and the second proteinase K. Diatoms were present in most organ samples of the immersed corpses; no diatoms could be found in the control samples. Our experience was that the enzymatic method seemed to be more convenient in terms of rapidity, safety and environmental protection than chemical digestion. The number of diatoms recovered with both methods was similar. Qualitative and quantitative analysis of both water and organ samples of immersion cases supported the diagnosis of death by drowning in 41.6% of the putrified cases studied. The authors suggest that diatom analysis using enzymatic digestion of organs can be used as a criterion for positive diagnosis of drowning in cases involving putrified bodies.     

Postmortem estimation of age at death based on aspartic acid racemization in dentin: Its applicability for root dentin

Summary The extent of aspartic acid racemization in total dentin and in dentin protein fractions from theroots of third molars was determined. In several cases coronal dentin was also investigated. The results of other authors, according to which the racemization of aspartic acid in root dentin apparently proceeds differently than in coronal dentin, could be confirmed. Consequently, the data published so far on age determination based on the extent of aspartic acid racemization in coronal dentin and the entire dentin of longitudinal sections cannot be applied to root dentin. In total root dentin and the acid soluble protein of root dentin, a close relationship was observed between the extent of aspartic acid racemization and age. Accordingly, estimation of age at death based on aspartic acid racemization in dentin is also possible for root dentin, apparently with good results. This is important particularly in those cases where a large portion of the coronal dentin is absent, for instance following dental treatment. In the investigation of root dentin, regression equations specific for root dentin must be employed in the estimation of age at death. Corresponding equations for third molars were calculated.     

Significance of detecting postmortem serum IgE in frozen corpses for the diagnosis of anaphylaxis in forensic

The aim of this study was to detect the postmortem serum total IgE levels in frozen corpses and identify whether the death incident caused by an anaphylaxis in forensic medicine. Autopsy cases with pathological death (total, n = 106; 4–214 h postmortem) include cardiac disease (n = 15), pulmonary infection (n = 12), central nervous system disorder (n = 6), pulmonary emboliszn (n = 7), hapetic disease (n = 5), kidney disease (n = 6), enteric disease (n = 10), necrotizing pancreatitis (n = 7), diffuse peritonitis (n = 6), MODS (n = 6), toxicosis (n = 5:), anaphylactic shock (n = 7), bronchial asthma (n = 8) and other disease (n = 6) were examined. Results showed that there was no significant difference between serum IgE levels and ages, postmortem intervals (PMIs), gender as well as survival time. Serum IgE levels of deaths due to anaphylactic shock and bronchial asthma were higher than that of other groups. Forensic pathology examination results showed the main pathology changes of bronchial asthma were mucous congestion in bronchial lumen and eosinophils infiltration in bronchial mucosa. The main pathological features of anaphylactic shock were laryngeal edema and eosinophils infiltration in multiple organs (lung and spleen). This research proved that there was a great significance for IgE to infer whether the individual died due to an anaphylaxis even for a long PMI in frozen corpses. Furthermore, we can also preliminarily determine the type of allergic death combined with the examination of forensic pathology. These findings further verify the feasibility of postmortem serum IgE in the diagnosis of forensic causes of death and broaden the application scope of this marker.     

Peri-mortem disease treatment: a little known cause of error in the estimation of the time since death in decomposing human remains

As part of a larger study into the estimation of the time since death of human bodies found decomposed, two bodies were monitored closely while decomposing in sequence in almost identical environmental situations. Both the rate and the manner, or stages, of decomposition that occurred to each body varied greatly and it is suggested that this was due to peri-mortem disease treatment of one of the bodies immediately prior to death. The results of this study suggest that certain forms of peri-mortem disease treatment are more important in modifying the process of post-mortem decomposition than previously thought to be case. In particular, cytotoxic drugs and/or antibiotic drug treatments in the peri-mortem period may contribute to altered rates and patterns of decomposition that have the potential to affect the estimation of the time since death in human bodies found decomposed.     

Post-mortem vitreous ammonium concentrations in estimating the time of death

Summary On a total of 150 specimens of vitreous humor the post-mortem ammonia value was determined by diffusion method. The results established that there existed a regular ammonia increase after a sudden death and that deviating high or low vitreous ammonia values are strongly correlated with the cause of death and with environmental temperature.The ammonia content of the vitreous humor extracted from both eyes at the same and at different post-mortem intervals showed no differences in the concentrations between both specimens sampled at the same moment while an obvious variation in the individual rate of the vitreous ammonia increase is found, especially when hospital cases with antemortem existing disease are concerned.

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Zusammenfassung Der Ammoniakgehalt der Glaskörperflüssigkeit wurde gemessen an insgesamt 150 Entnahmen mit bekannter Todeszeit nach der Diffusionsmethode. Die Ergebnisse zeigen einen regelmässigen postmortalen Anstieg des Ammoniaks in plötzlichen Todesfällen während abweichend höhere und niedrigere Messzahlen sich eng korreliert verhalten mit der Todesursache und dem Aussentemperatur. Doppelbestimmungen des Ammoniakgehalts im Corpus Vitreum beider Augen zu gleicher und verschiedener Leichenzeit vorgenommen erweisen keine Konzentrationsunterschiede in den zu gleicher Zeit entnommenen Proben aber eine merkbare Schwankung in dem individuellen Anstieg der Ammoniakwerte des Glaskörpers, besonders wenn Hospitalfälle mit vorherigen Krankheiten betroffen sind. Wegen der Beschränkung einer notwendigen Selektion in bezug auf Ursache und Plötzlichkeit eines Sterbens ist die Methodik der Ammoniakbestimmung im Glaskörperinhalt angesichts der mit zunehmender Liegezeit wachsenden individuellen Streuung nur als zusätzliches Hilfsmittel zur Todeszeitschätzung verwendbar.

     

Experimental studies on the kinetics of early postmortem decay of the lymphocyte membrane

Summary Tests on human and animal cadavers were carried out to investigate the early postmortem decay of the lymphocyte membrane.HLA-typing of cool-kept human cadavers can be performed within the first 36 h after death. Hereby, a possibility is obtained to determine HLA-ABC antigens of a deceased for hemogenetic or cornea transplantation purposes. The data obtained from animal experiments give evidence that the decay of the lymphocyte membrane follows a function of the Verhult's type. Using this function, it is possible to estimate the rate constant as a function of temperature.Partially presented to the German Society of Forensic Medicine at a meeting in Heidelberg, and to the Working Group for Histocompatibility Testing at a meeting in Freiburg, both in 1980     

Medical death certification by forensic physicians in the Netherlands: Validity and interdoctorvariation

The aim of the study was to assess interdoctorvariation and validity in death certification by forensic physicians using 19 written scenarios. The scenarios described typical cases from forensic-medical practice. Physicians were asked to determine the manner of death (natural/unnatural) and to provide an ICD-10 code for the cause of death. In contrast to most studies on this topic, the measure of agreement among physicians was chance-corrected and a standard was used to assess the correctness of the assigned cause and manner of death. Forty-seven physicians participated in the survey. The study demonstrated that forensic physicians varied widely in their conclusions. With respect to manner of death, adequate agreement (defined as kappa>0.70) was achieved in six scenarios (32% of all scenarios). Concerning the underlying cause of death, adequate agreement was reached in three cases (16% of all scenarios). Furthermore, predictors for the correctness of manner and cause of death were studied using logistic regression. Years of experience as a forensic physician significantly predicted the correctness of cause of death (p < 0.05). Other predictors remained insignificant. With regard to manner of death, none of the studied predictors proved to be significant. To conclude, there appears to be a lack of consistency among forensic physicians regarding death certification. The ICD-10 coding of causes of death applied by forensic physicians is questionable. Less experienced physicians need supervision by more experienced colleagues when making judgments concerning the cause of death. Altogether, there is an urgent need to work out consensus-based guidelines for forensic physicians on how to certify deaths.     

A method for estimating time since death through analysis of substances deposited on the surface of dental enamel in a body immersed in freshwater

Estimating the elapsed time after death of a corpse immersed in freshwater is often challenging owing to body temperature changes and the degree of postmortem lividity, and the tissues used for this assessment deteriorate drastically as the postmortem period progresses. In this study, assuming a corpse was immersed in freshwater, we analyzed changes in the amount of deposits on the enamel surface of teeth since the time of immersion using an electron probe microanalyzer. We calculated a regression equation for estimating the time of immersion in water (the time after death). We considered this approach would be useful to estimate the time after death even in rotting and skeletonized corpses from which only limited information could be obtained. This study was undertaken according to a previously reported method that used a regression equation for estimating the time after death of bodies found in a seawater area.     

Correlation between the post-mortem cell content of cerebrospinal fluid and time of death

Summary The number of cells in the cerebrospinal fluid increases after death. It is not clear whether this observation represents a post-mortem or a supravital phenomenon. There is speculation that some of the cells actively enter the cerebrospinal fluid during the first hours after death and that the rest exfoliate from the subarachnoid layer. The post-mortem cell count in cerebrospinal fluid correlates to the time after death and can be described mathematically (polynomial curve of third order).     

Determining time of death using blow fly eggs in the early postmortem interval

A forensic entomology case is presented which is interesting for two reasons; firstly, it uses egg development, but secondly, it involves a case which is over 20 years old, and was recently re-opened. The only entomological evidence was in the form of photographs. Usually, it is not possible to make any determinations from photographs alone, as species and age cannot be determined. However, this case was particularly unusual, as close up photographs taken by police at the crime scene showed the first egg eclosion. Weather records, developmental data and degree day accumulations, together with a knowledge of local species and their habitats, allowed determination of the time since death. Time of death was affirmed when the defendant was convicted of first degree murder more than 20 years after the killing.     

Exchanged identities in a complex multiple homicide case. Identification and cause of death

We report a multiple homicide in Angola involving six Portuguese citizens. Immediately after the crime, four bodies were retrieved from the homicide site. Forensic autopsies were allegedly performed by local doctors and three bodies were transferred to Portugal, where they were buried with no extra expertise, presumably identified only by the examinations carried out in Angola. The two remaining bodies, presumably from sub-adults, were only discovered 1 year later. At that time, some bone samples of these two corpses were sent to Portugal. A multidisciplinary team of the National Institute of Legal Medicine (NILM) in Portugal, including forensic anthropologist and pathologists, and forensic geneticist, found out that the bodies were previously misidentified. In fact, the anthropological and genetic examinations on the remains of the two persons showed that they did not belong to a child and an adolescent but instead to two adult victims, at least one of them was supposed to be buried in Portugal since 1 year. The verification of this misunderstanding led to a series of exhumations, of the three victims previously buried in Portugal. In all, the identities were reconstructed and the cause of death could be established in four of them. A multiplicity of severe traumatic cranial injuries were detected, which were, undoubtedly due to a violent death.