Genotyping strategies for tissues fixed with various embalming fluids for human identification, databasing, and traceability.

Madeline Ashton, Natalia Czado, Michelle Harrel, Sheree Hughes
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Abstract

Within anatomical willed body programs and skeletal collections, whole bodies and their disassociated limbs and organs are identified and tracked. However, if these tracking mechanisms fail, DNA recovered from the formalin-fixed tissues/organs could provide an additional layer of quality assurance. Embalming fluids preserve biological tissues; however, they also damage, fragment, and cross-link DNA and protein molecules. This project investigated the success of STR-typing from various soft tissue and bone samples that were fixed with embalming solutions with a range of formaldehyde concentrations. Formalin-fixed samples dissected from five cadavers, including skin, muscle, bone, heart, and kidney were used in Phase 1 of this study. In Phase 2, an additional 57 tissue samples from various embalmed organs and body parts were collected to demonstrate long-term fixation and direct applicability within a body donor program. DNA was extracted from the samples using the QIAamp® FFPE Tissue Kit (QIAGEN), quantified with the Investigator® QuantiPlex® Pro RGQ qPCR Kit (QIAGEN), and amplified using the Investigator® 24plex and 26plex QS Kits and the Investigator® DIPplex Kit (QIAGEN). The results show the DNA was severely damaged, degraded, and often in low amounts (after one year post-embalming). Sampling from skin and muscle tissues embalmed with ~2.5%-5% formaldehyde solutions appears to be the best strategy for identification, while also maintaining the preservation of the tissues. The results of this project can provide informative data when determining which genotyping strategy may be best suited for the identification, re-association, and establishment of a database for the provenance of formalin-fixed human remains.

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用各种防腐液固定的组织的基因分型策略,用于人类识别、数据库和可追溯性。
在解剖意愿身体程序和骨骼采集中,整个身体及其分离的肢体和器官都被识别和跟踪。然而,如果这些跟踪机制失败,从福尔马林固定的组织/器官中回收的DNA可以提供额外的质量保证层。防腐液保存生物组织;然而,它们也会损伤、碎片化和交联DNA和蛋白质分子。该项目调查了用一系列甲醛浓度的防腐溶液固定的各种软组织和骨骼样本的STR分型的成功。本研究的第一阶段使用了从五具尸体上解剖的福尔马林固定样本,包括皮肤、肌肉、骨骼、心脏和肾脏。在第二阶段,从各种防腐器官和身体部位收集了另外57个组织样本,以证明长期固定和在身体捐献者计划中的直接适用性。使用QIAamp®FFPE组织试剂盒(QIAGEN)从样本中提取DNA,使用Investigator®QuantiPlex®Pro RGQ qPCR试剂盒(QIAGEN)进行定量,并使用Investirator®24plex和26plex QS试剂盒以及Investigator®;DIPplex试剂盒(Q IAGEN)进行扩增。结果表明,DNA严重受损、降解,而且数量通常很少(防腐一年后)。从用~2.5%-5%甲醛溶液防腐的皮肤和肌肉组织中取样似乎是鉴定的最佳策略,同时也能保持组织的保存。该项目的结果可以在确定哪种基因分型策略最适合福尔马林固定人类遗骸来源的鉴定、重新关联和建立数据库时提供信息性数据。
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