识别体液和器官组织非常重要,因为它们可以在刑事调查中提供关键证据,并帮助法院做出明智的决定。主要通过评估生物来源,并可能在活动水平直至死亡或致命损害。在这项研究中,从Illumina的9个器官组织的甲基化EPIC阵列数据中鉴定出器官组织特异性CpG标记,包括表皮,真皮,心,骨骼肌,血,肾,大脑,肺,还有肝脏,来自10名韩国人的尸检。通过使用43个样本的验证测试,18个低甲基化标记,每种器官组织类型都有两个标记,选择构建SNaPshot测定。设计了涉及正向和反向SBE引物的两个多重测定,以帮助研究者通过重复分析相同的PCR产物的标记来准确地确定所分析的组织样品的器官来源。开发的多路复用显示出高精度,在来自10名亚洲人的尸检的88个样本中,对9种器官组织类型的存在实现100.0%的正确检测。然而,两个肺样本显示血液存在的其他阳性迹象.使用80个尸检样本(心脏,骨骼肌,血,肾皮质,肾髓质,大脑,肺,和肝脏)来自德国的10名个体,显示出总体可比较的结果,正确检测了92.5%样品(80个样品中的74个)中8种器官组织类型的存在。在六个样本的情况下,由于靶组织标志物位点的未甲基化信号缺失,因此无法成功确定正确的组织.这些肺样品中的一个仅显示血液的非预期脱靶信号。观察到的差异可能是由于常规尸检期间样本收集的差异,由于PCR循环仪的技术差异,以及用于信号呼叫的阈值。通过应用更严格的低甲基化阈值,表明存在其他组织类型和脱靶非甲基化信号似乎可以缓解。因此,开发的SNaPshot多重分析对于处理器官组织鉴定的法医调查人员将是有价值的,以及旨在确定犯罪现场情况的检察官和辩方。
Identifying body fluids and organ tissues is highly significant as they can offer crucial evidence in criminal investigations and aid the court in making informed decisions, primarily through evaluating the biological source and possibly at the activity level up to death or fatal damage. In this study, organ tissue-specific CpG markers were identified from Illumina\'s methylation EPIC array data of nine organ tissues, including epidermis, dermis, heart, skeletal muscle, blood, kidney, brain, lung, and liver, from autopsies of 10 Koreans. Through the validation test using 43 samples, 18 hypomethylation markers, with two markers for each organ tissue type, were selected to construct a
SNaPshot assay. Two multiplex assays involving forward and reverse SBE primers were designed to help investigators accurately determine the organ origin of the analyzed tissue samples through repeated analysis of the same PCR products for markers. The developed multiplex demonstrated high accuracy, achieving 100.0 % correct detection of the presence of nine organ tissue types in 88 samples from autopsies of 10 Asians. However, two lung samples showed additional positive indications of the presence of blood. An interlaboratory comparison using 80 autopsy samples (heart, skeletal muscle, blood, kidney cortex, kidney medulla, brain, lung, and liver) from 10 individuals in Germany revealed overall comparable results with correct detection of the presence of eight organ tissue types in 92.5 % samples (74 of 80 samples). In the case of six samples, it was impossible to determine the correct tissue successfully due to drop-outs of unmethylation signals at target tissue marker loci. One of these lung samples revealed only non-intended off-target signals for blood. The observed differences might be due to differences in sample collection during routine autopsy, technical differences due to the PCR cycler, and the threshold used for signal calling. Indicating the presence of additional tissue type and off-target unmethylation signals seems alleviated by applying more stringent hypomethylation thresholds. Therefore, the developed
SNaPshot multiplex assays will be valuable for forensic investigators dealing with organ tissue identification, as well as for prosecutors and defense aiming to establish the circumstances that occurred at the crime scene.