Forensic application of three interstitial pneumonia markers: search for new pneumonia markers in dead bodies.

In forensic cases, detailed identification of pneumonia is important. Our objective was to statistically determine the applicability of three interstitial lung disease (ILD) markers for forensic diagnosis using serum collected from dead bodies with various postmortem intervals (PMIs). We retrospectively analyzed the levels of postmortem serum Krebs von den Lungen-6 (KL-6) and pulmonary surfactant-associated proteins A and D (SP-A and SP-D) using 221 samples obtained during forensic autopsy at our facility from 2019 to 2023. We evaluated the diagnostic efficacy of ILD markers for various pneumonias against the pathological diagnosis, and examined the assessment of the severity of ILD. When comparing the ILD group with bacterial pneumonia (BP) versus the control group, there was a significant increase in KL-6 in the ILD group. When comparing the severe ILD (SILD) group with the mild ILD (MILD) group, there was a significant increase in KL-6 and SP-D in the SILD group. The optimal cutoff values for differentiating SILD were 607.0 U/mL for KL-6, 55.5 ng/mL for SP-A, and 160.0 ng/mL for SP-D, and the sensitivity/specificity (%) of KL-6, SP-A, and SP-D for SILD were 84.1/95.2, 55.6/85.7, and 66.7/74.6, respectively. This is the first study to examine KL-6 in postmortem serum in forensic medicine. By analyzing dead bodies with various PMIs, our results confirmed statistically that postmortem serum KL-6 specifically detects ILD, postmortem serum SP-A has high sensitivity to lung injury, and postmortem serum SP-D is potentially useful in assessing the severity of ILD.

Sex estimation from skull measurements of a contemporary Japanese population using three-dimensional computed tomography images.

In this study, we assessed the sexual dimorphism of the contemporary Japanese skull and established sex discriminant function equations based on cranial measurements using three-dimensional (3D) computed tomography (CT) images. The CT images of 263 corpses (142 males, 121 females) that underwent postmortem CT scanning and subsequent forensic autopsy were evaluated. Twenty-one cranial measurements were obtained from 3D CT reconstructed images, which extracted only bone data. We performed descriptive statistics and discriminant function analyses for the measurements. Nineteen measurements were significantly larger in males, suggesting sexual dimorphism of the Japanese skulls. Univariate discriminant function analyses using these measurements showed a sex classification accuracy of 57.8-88.2%, and bizygomatic breadth provided the highest correct prediction rate. Multivariate discriminant function analyses offered the most accurate model using seven variables with an estimation rate of 93.9%. Our results suggest that cranial measurements based on 3D CT images may help in the sex estimation of unidentified bodies in a contemporary Japanese population.

Genomic analysis of SARS-CoV-2 in forensic autopsy cases of COVID-19.

Numerous genomic analyses of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been conducted, highlighting its variations and lineage transitions. Despite the importance of forensic autopsy in investigating deaths due to coronavirus disease 2019 (COVID-19), including out-of-hospital deaths, viral genomic analysis has rarely been reported due in part to postmortem changes. In this study, various specimens were collected from 18 forensic autopsy cases with SARS-CoV-2 infection. Reverse-transcription quantitative polymerase chain reaction revealed the distribution of the virus in the body, primarily in the respiratory organs. Next-generation sequencing determined the complete genome sequences in 15 of the 18 cases, although some cases showed severe postmortem changes or degradation of tissue RNA. Intrahost genomic diversity of the virus was identified in one case of death due to COVID-19. The accumulation of single-nucleotide variations in the lung of the case suggested the intrahost evolution of SARS-CoV-2. Lung of the case showed diffuse alveolar damage histologically and positivity for SARS-CoV-2 by immunohistochemical analysis and in situ hybridization, indicating virus-associated pneumonia. This study provides insights into the feasibility of genomic analysis of SARS-CoV-2 in forensic autopsy cases and the potential for uncovering important information in COVID-19 deaths, including out-of-hospital deaths.

Stature estimation by semi-automatic measurements of 3D CT images of the femur.

Stature estimation is one of the most basic and important methods of personal identification. The long bones of the limbs provide the most accurate stature estimation, with the femur being one of the most useful. In all the previously reported methods of stature estimation using computed tomography (CT) images of the femur, laborious manual measurement was necessary. A semi-automatic bone measuring method can simplify this process, so we firstly reported a stature estimation process using semi-automatic bone measurement software equipped with artificial intelligence. Multiple measurements of femurs of adult Japanese cadavers were performed using automatic three-dimensional reconstructed CT images of femurs. After manually setting four points on the femur, an automatic measurement was acquired. The relationships between stature and five femoral measurements, with acceptable intraobserver and interobserver errors, were analyzed with single regression analysis using the standard error of the estimate (SEE) and the coefficient of determination (R2). The maximum length of the femur (MLF) provided the lowest SEE and the highest R2; the SEE and R2 in all cadavers, males and females, respectively, were 3.913 cm (R2 = 0.842), 3.664 cm (R2 = 0.705), and 3.456 cm (R2 = 0.686) for MLF on the right femur, and 3.837 cm (R2 = 0.848), 3.667 cm (R2 = 0.705), and 3.384 cm (R2 = 0.699) for MLF on the left femur. These results were non-inferior to those of previous reports regarding stature estimation using the MLF. Stature estimation with this simple and time-saving method would be useful in forensic medical practice.

Trends and Characteristics in Health Care-related Deaths Investigated through Medico-legal Autopsies after System Changes in Japan.

In Japan, a new cause-of-death investigation system and related new laws were enacted in the mid-2010s. These laws provided for an autopsy system for non-criminal unnatural deaths and a medical accident investigation system outside the criminal justice process for health care-related deaths. We retrospectively explored changes in the number and characteristics of medico-legal autopsy cases of health care-related deaths in Chiba Prefecture, Japan, and examined trends over time during these reforms. We found that the percentage of forensic autopsies based on the Code of Criminal Procedure for health care-related deaths had decreased significantly. The number of autopsies of accidental and unintentional deaths in nursing homes, which are not covered by the newly established medical accident investigation system, has been increasing, reflecting the ageing of society. The trend toward decriminalisation of health care-related deaths was expected to contribute more to medical safety if the scope was expanded and a system for disclosure of autopsy information was established.

Postmortem CT and MRI findings of massive fat embolism.

OBJECTIVE: To elucidate postmortem computed tomography (PMCT) and postmortem magnetic resonance (PMMR) imaging findings suggesting massive fat embolism. MATERIALS AND METHODS: Consecutive forensic cases with PMCT and PMMR scans of subjects prior to autopsy were assessed. For PMCT, 16- or 64-row multidetector CT scans were used; for PMMR, a 1.5 T system was used. MRI sequences of the chest area included T2- and T1-weighted fast spin-echo imaging, T2*-weighted imaging, T1-weighted 3-dimensional gradient-echo imaging with or without a fat-suppression pulse, short tau inversion recovery, and in-phase/opposed-phase imaging. At autopsy, forensic pathologists checked for pulmonary fat embolism with fat staining; Falzi's grading system was used for classification. RESULTS: Of 31 subjects, four were excluded because fat staining for histopathological examination of the lung tissue could not be performed. In three of the remaining 27 subjects, histology revealed massive fat embolism (Falzi grade III) and the cause of death was considered to be associated with fat embolism. CT detected a "fat-fluid level" in the right heart or intraluminal fat in the pulmonary arterial branches in two subjects. MRI detected these findings more clearly in both subjects. In one subject, CT and MRI were both negative. There were no positive findings in the 24 subjects that were fat embolism-negative by histology. DISCUSSION AND CONCLUSION: In some subjects, a massive fat embolism can be suggested by postmortem imaging with a "fat-fluid level" in the right heart or intraluminal fat in the pulmonary arterial branches. PMMR potentially suggests fat embolism more clearly than PMCT.

Fatal hemorrhage complicated with methamphetamine poisoning and its post-mortem CT features.

The purpose of this study was to assess the incidence of fatal hemorrhage complicated with methamphetamine (MA) poisoning and to examine the postmortem computed tomography (PMCT) features of fatal intracerebral hemorrhage (ICH) with and without MA poisoning. The study also attempted to determine the differences in PMCT between those two groups. Consecutive medicolegal autopsy data from November 2011 to February 2018 were searched for 3044 cases. First, the incidence and distribution of all cases of nontraumatic fatal hemorrhage with various causes were examined. Second, cases of ICH on the basal ganglia and brain stem were extracted. The PMCT findings were compared with respect to nine parameters: volume of hematoma, ventricular perforation, midline shift distance, aortic calcification, calcification of aortic valve, calcification of coronary artery, cardiothoracic ratio, circumference of ascending aorta, and volume of bladder contents. Of the 3044 cases, 97 were nontraumatic fatal hemorrhage; of these 97 cases, 20 were classified as MA poisoning with 9 ICH cases, and 60 cases were classified as non-MA poisoning with 14 ICH cases. A statistically significant difference in ages was observed between the two groups. On PMCT comparison of ICH, statistically significant differences were evident in the midline shift distance and calcification of the aortic valve. Forensic radiologists should be aware of the possibility of ICH with MA poisoning if fatal hemorrhage is detected on PMCT. Younger age, less calcification of the aortic valve, and a remarkable midline shift may be the keys to recognition.

Postmortem angiography revealing traumatic rupture of the intracranial internal carotid artery.

Diagnosis of traumatic subarachnoid hemorrhage (SAH), although relatively rare, is important in forensic medicine. It is mostly associated with rupture of the vertebrobasilar artery. Traumatic aneurysm of the intracranial part of the internal carotid artery (ICA) is also rare but has been reported in several studies. It is thought that the intracranial ICA is injured by blunt force to the head, neck, and chest. However, traumatic SAH with fatal acute course resulting from rupture of the ICA is especially uncommon: only two fatal cases without an associated aneurysm have been reported in the English-language literature. Although detecting the arterial lesion is required to make a precise diagnosis, this is sometimes impossible by macroscopic examination at autopsy or by investigation after formalin-fixation according to the position of the lesion. We report a rare case of fatal traumatic SAH associated with intracranial ICA rupture. Postmortem computed tomography angiography was useful to confirm the lesion.

Determination of sex on the basis of hyoid bone measurements in a Japanese population using multidetector computed tomography.

Sex determination for unidentified human remains is important because the determined sex is usually used as a foundation for the estimation of other biological profiles. This study investigated the sexual dimorphism of the adult hyoid bone and developed discriminant equations to accurately estimate sex in a contemporary Japanese population using measurements on two-dimensional computed tomography (CT) images. The study sample comprised 280 cadavers (140 males, 140 females) of known age and sex that underwent postmortem CT and subsequent forensic autopsy. In accordance with previous studies, seven measurements were performed using two-dimensional CT reconstructed images. Most measurements of males were significantly greater than those of females. The perpendicular length from the most anterior edge of the hyoid body to the line that connects the most distal points of the greater horns of the hyoid and the linear distance between the most lateral edges of the hyoid body most significantly contributed to sex determination. This study demonstrated that discriminant functions on the basis of three measurements provided higher rates of accurate sex classification (93.3-94.6%) than univariate functions. Therefore, the hyoid bone is highly sexually dimorphic in the contemporary Japanese population and may be useful in forensic contexts for sex determination with a high level of accuracy.

Stature estimation based on femoral measurements in the modern Japanese population: a cadaveric study using multidetector computed tomography.

We aimed to reproduce the anthropometrical measurement of femoral dimensions using multi-planar reconstruction computed tomography (CT), assess the correlation between stature and femoral measurements obtained by this approach, and establish a regression equation for estimating stature in the modern Japanese population. We used data regarding 224 cadavers (116 males, 108 females) that were subjected to postmortem CT and subsequent forensic autopsy at our department between October 2009 and July 2016. To simulate the placement of the femur on the osteometric board using reconstructed CT images, we defined a virtual horizontal plane (VHP) based on the three most dorsal points of the femur (lateral condyle, medial condyle, and greater trochanter). Five femoral measurements including the maximum femoral length (MFL) were obtained. The correlations between stature and each femoral measurement were expressed in terms of the coefficient of determination (R2). On regression analysis, MFL provided the lowest value for the standard error of the estimation (SEE); the SEE values in all subjects, males, and females, respectively, were 3.783 cm (R2 = 0.832), 3.850 cm (R2 = 0.653), and 3.340 cm (R2 = 0.760) for MFL on the left side and 3.747 cm (R2 = 0.835), 3.847 cm (R2 = 0.650), and 3.290 cm (R2 = 0.687) for MFL on the right side. Multiple regression equations using MFL and femoral epicondylar breadth were slightly superior to simple regression equations in males and in all subjects (SEE = 3.44-3.55 cm), whereas no effective equation could be obtained in females. To our knowledge, this is the first multiple regression equation for stature estimation using only femoral measurements.

Myoglobinemia markers with potential applications in forensic sample analysis: lipid markers in myoglobinemia for postmortem blood.

The crush syndrome, in which rhabdomyolysis and trauma occur as a result of heat stroke and drug intoxication, can lead to myoglobinemia. This condition can be diagnosed by measuring myoglobin (Mb) levels in blood and urine. However, postmortem Mb levels are unreliable indicators, since blood Mb concentration drastically increases within a very short time after death and urine cannot always be obtained at dissection; this makes it difficult to diagnose myoglobinemia in a corpse. To address this issue, in this study, we used a lipidomics approach to identify markers that can be used to detect myoglobinemia in postmortem blood samples. We found that increases in levels of fatty acid oxides such as stearic, oleic, linoleic, and arachidonic acid and decreases in levels of plasmalogens and phosphatidylethanolamine in the blood were associated with high Mb level. These results demonstrate that postmortem samples are amenable to lipidomics analysis and provide a set of markers other than Mb that can be used for postmortem diagnosis of myoglobinemia.

Frequency and influencing factors of cardiopulmonary resuscitation-related injuries during implementation of the American Heart Association 2010 Guidelines: a retrospective study based on autopsy and postmortem computed tomography.

AIM: To determine the frequency of cardiopulmonary resuscitation (CPR)-related injuries and factors involved in their occurrence, data based on forensic autopsy and postmortem computed tomography (PMCT) during implementation of the 2010 American Heart Association Guidelines for CPR were studied. METHODS: We retrospectively evaluated data on adult patients with non-traumatic deaths who had undergone manual CPR and autopsy from January 2012 to December 2014. CPR-related injuries were analyzed on autopsy records and PMCT images and compared with results of previous studies. RESULTS: In total, 180 consecutive cases were analyzed. Rib fractures and sternal fractures were most frequent (overall frequency, 66.1 and 52.8%, respectively), followed by heart injuries (12.8%) and abdominal visceral injuries (2.2%). Urgently life-threatening injuries were rare (2.8%). Older age was an independent risk factor for rib fracture [adjusted odds ratio (AOR), 1.06; 95% confidence interval (CI), 1.04-1.08; p < 0.001], ≥ 3 rib fractures (AOR, 1.06; 95% CI, 1.02-1.09; p = 0.002), and sternal fracture (AOR, 1.03; 95% CI, 1.01-1.05; p < 0.001). Female sex was significantly associated with sternal fracture (AOR, 2.08; 95% CI, 1.02-4.25; p = 0.04). Chest compression only by laypersons was inversely associated with rib and sternal fractures. Body mass index and in-hospital cardiac arrest were not significantly associated with any complications. The frequency of thoracic skeletal injuries was similar to that in recent autopsy-based studies. CONCLUSIONS: Implementation of the 2010 Guidelines had little impact on the frequency of CPR-related thoracic skeletal injuries or urgently life-threatening complications. Older age was the only independent factor related to thoracic skeletal injuries.

An unusual autopsy case of lethal hypothermia exacerbated by body lice-induced severe anemia.

Pediculus humanus humanus (known as body lice) are commonly found in the folds of clothes, and can cause skin disorders when they feed on human blood, resulting in an itching sensation. Body lice are known as vectors of infectious diseases, including typhus, recurrent fever, and trench fever. An infestation with blood-sucking body lice induces severe cutaneous pruritus, and this skin disorder is known as "vagabond's disease." A body lice infestation is sometimes complicated with iron deficiency anemia. In the present case, a man in his late 70s died of lethal hypothermia in the outdoors during the winter season. The case history and autopsy findings revealed that the cause of the lethal hypothermia was iron deficiency anemia, which was associated with a prolonged infestation of blood-sucking body lice. Also, he had vagabond's disease because the skin on his body was abnormal and highly pigmented. This is an unusual autopsy case since the body lice contributed to the cause of the death.

A case of acute subdural hematoma due to ruptured aneurysm detected by postmortem angiography.

Acute subdural hematoma (ASDH) is mostly caused by head trauma, but intrinsic causes also exist such as aneurysm rupture. We describe here a case involving a man in his 70s who was found lying on the bedroom floor by his family. CT performed at the hospital showed ASDH and a forensic autopsy was requested. Postmortem cerebral angiography showed dilatation of the bifurcation of the middle cerebral artery, which coincided with the dilated part of the Sylvian fissure. Extravasation of contrast medium into the subdural hematoma from this site was suggestive of a ruptured aneurysm. Autopsy revealed a fleshy hematoma (total weight 110 g) in the right subdural space and findings of brain herniation. As indicated on angiography, a ruptured saccular aneurysm was confirmed at the bifurcation of the middle cerebral artery. Obvious injuries to the head or face could not be detected on either external or internal examination, and intrinsic ASDH due to a ruptured middle cerebral artery aneurysm was determined as the cause of death. One of the key points of forensic diagnosis is the strict differentiation between intrinsic and extrinsic onset for conditions leading to death. Although most subdural hematomas (SDH) are caused by extrinsic factors, forensic pathologists should consider the possibility of intrinsic SDH. In addition, postmortem angiography can be useful for identifying vascular lesions in such cases.

Evaluations of lipid peroxidation and inflammation in short-term glycerol-induced acute kidney injury in rats.

Rhabdomyolysis is characterised by acute kidney injury (AKI) resulting from skeletal muscle injury. Lipid peroxidation-mediated oxidant injury and pro-inflammatory cytokine-mediated inflammatory response play critical roles in the pathogenesis of rhabdomyolysis-induced AKI. The present study aimed to investigate the short-term effects of both lipid peroxidation and inflammatory responses on rhabdomyolysis-induced AKI in a rat model of glycerol-induced rhabdomyolysis. Rhabdomyolysis was induced by the intramuscular injection of 50% glycerol in saline (10 mL/kg) into the hind limbs of rats. Rats were killed 1 or 3 hours after glycerol injection. Time-dependent increases in serum biochemical parameters, including blood urea nitrogen, creatinine, lactate dehydrogenase and creatine phosphokinase levels, were observed 1 hour after glycerol injection. In kidneys, glycerol injection resulted in histopathological changes such as renal tubular injury and renal tubular myoglobin deposition. Levels of Nε-(hexanoyl)lysine-modified, 4-hydroxy-2-nonenal-modified, and nitrotyrosine-modified proteins in rat kidneys were unaltered at 1 hour after glycerol injection, but increased significantly at 3 hours. Increases in renal nitric oxide production and the expression levels of inducible nitric oxide synthase, interleukin-6 and tumour necrosis factor-α in the renal parenchyma were observed at 1 hour after glycerol injection and plateaued at 3 hours. Our findings suggest that the pro-inflammatory cytokine-mediated inflammatory response may cause rhabdomyolysis-induced AKI very shortly after glycerol injection, and lipid peroxidation-mediated oxidant injury may promote the development of these pathophysiological processes.

Stature estimation in Japanese cadavers based on pelvic measurements in three-dimensional multidetector computed tomographic images.

The aim of this study was to examine the feasibility of stature estimation by measuring the pelvic bones of Japanese cadavers using three-dimensional (3D) computed tomography (CT). We assessed 3D reconstructed images of 210 Japanese subjects (108 males, 102 females) who had undergone postmortem CT between May 2011 and November 2013. We defined the linear distance from the anterosuperior margin of the left and right anterior superior iliac spines (ASIS) to the posterior margin of the left and right ischial spines as the LSS and RSS, respectively. We also defined the linear distance from the anterosuperior margin of the left and right ASIS to the anteroinferior margin of the left and right ischial tuberosities as the LST and RST, respectively. The correlation between the cadaver stature (CS) and each parameter (LSS, LST, RSS, and RST) was evaluated using Pearson product-moment correlation coefficients and regression analysis was performed for stature estimation. All four parameters correlated significantly with stature independent of sex, suggesting that they can be used as a tool for stature estimation. The LST had the closest correlation with stature in both sexes.

Fluid-Fluid Level and Pericardial Hyperdense Ring Appearance Findings on Unenhanced Postmortem CT Can Differentiate Between Postmortem and Antemortem Pericardial Hemorrhage.

OBJECTIVE: The objective of this study was to determine whether selected postmortem unenhanced CT findings can discriminate between antemortem and postmortem pericardial hemorrhage. MATERIALS AND METHODS: Thirty-one consecutive cases of postmortem CT followed by autopsy identifying at least 50 mL of hemorrhaged blood were reviewed. Seven cases were classified as postmortem pericardial hemorrhage secondary to chest compression (postmortem group), and 24 cases were classified as antemortem pericardial hemorrhage secondary to disease or trauma (antemortem group), on the basis of autopsy findings. Postmortem CT findings of pericardial hemorrhage were classified as stratification comprising upper low-density and lower high-density areas (i.e., fluid-fluid level) and a high-density concentric ring (i.e., pericardial hyperdense ring). Diagnostic values for detecting antemortem or postmortem pericardial hemorrhage using this classification system, along with attenuation (in Hounsfield units) of pericardial hemorrhage lesions and the presence or absence of flattened heart sign on postmortem CT, were measured. RESULTS: There were statistically significant differences in fluid-fluid level and pericardial hyperdense ring between the postmortem and antemortem groups (p < 0.001). The sensitivity, specificity, positive predictive value, and negative predictive value of fluid-fluid level for detecting postmortem pericardial hemorrhage were 86%, 96%, 86%, and 100%, respectively. The mean attenuation of pericardial hemorrhage lesions differed statistically significantly between the groups (p = 0.004). The presentation of the flattened heart sign did not differ statistically significantly between the groups (p = 0.681). CONCLUSION: Fluid-fluid level and pericardial hyperdense ring on postmortem CT, combined with attenuation, are useful for differentiating between antemortem and postmortem pericardial hemorrhage secondary to chest compression.

Observation of vertebral artery damage using angioscopy in autopsy cases.

The vertebral arteries are important blood vessels that supply the cerebral circulation in conjunction with the internal carotid arteries. In cases of subarachnoid hemorrhage, it is necessary to examine the vertebral arteries as potential sources of bleeding due to blunt trauma (head and neck) or of cerebral embolism that originated on the surface of the damaged intima as a result of hyperflexion or hyperextension. However, a considerable part of the vertebral arterial surface is surrounded by bone, resulting in challenges during examination in a routine autopsy. In this study, angioscopy was used to inspect the vertebral artery intima for damage in cases of neck injury, head injury, or neck strangulation. Intimal damage was detected in 34 out of the total 75 cases. Of the 28 cases with cervical discopathy or fracture, 61% had intimal damage. In addition, postmortem application of computed tomography angiography was performed to identify the injured vessel in a case with traumatic subarachnoid hemorrhage, and a perforated hole was detected using angioscopy, which did not introduce autopsy-related artifacts. Therefore, angioscopy may be a useful and nondestructive method to identify intimal damage in the vertebral arteries during an autopsy.

Age estimation by quantitative features of pubic symphysis using multidetector computed tomography.

Macroscopic assessment of the pubic symphysis is commonly used for age estimation because its surface changes over time. However, postmortem computed tomography (PMCT), a method several forensic medical departments and institutes have begun to adopt, has the potential to simplify the information gathering process from the pelvic bone without requiring soft tissue removal. Some studies have previously evaluated the use of three-dimensional images of the pubic symphysis, but because of variance in the graphics processing among image analysis software packages, certain differences have been observed between these studies. Therefore, in this study, the PMCT findings of 199 subjects of known age and sex were retrospectively reviewed to examine the feasibility of age estimation using planar images of the pubic bones and soft tissue. The coronal and axial sectional images were observed at the center of the symphyseal surface, and the pubic bone length and thickness of the connective tissue of the pubic symphysis were measured at each slice. Our results revealed a significant positive correlation between the length of the pubic bone of the coronal section and age, suggesting that the use of a cutoff value for pubic bone length might be feasible for age estimations. In addition, the thickness of the connective tissue tended to narrow over time. Although the prediction interval range of planar images obtained by PMCT was major and is not usable in practice at this moment, it may still be a useful tool if used in conjunction with other findings obtained by PMCT.

Genotyping of 38 insertion/deletion polymorphisms for human identification using universal fluorescent PCR.

Short insertion/deletion (Indel) polymorphisms of approximately 2-6 bp are useful as biallelic markers for forensic analysis, and the application of Indel genotyping as a supplementary tool would improve human identification accuracy. We examined the allele frequencies of 37 autosomal Indels in the Japanese population and developed a novel dual-color genotyping method for human identification on the basis of universal fluorescent PCR, including the sex-typing amelogenin locus. Target genomic fragment sizes for 38 Indels were 49-143 bp. We analyzed these Indels in 100 Japanese individuals using the M13(-47) sequence as a universal primer. For dual-color genotyping, we designed a novel universal primer with high amplification efficiency and specificity. Using FAM-labeled M13(-47) and HEX-labeled modified M13(-47) primers, fluorescent signals at all loci were clearly distinguished in two independent multiplex PCRs. Average minor allele frequency was 0.39, and accumulated matching probability was 2.12 × 10(-15). Complete profiles were successfully amplified with as little as 0.25 ng of DNA. This method provides robust, sensitive, and cost-effective genotyping for human identification.