Dutta Ranjan, Mahajan Kedar R, Nakamura Kunio, Ontaneda Daniel, Chen Jacqueline, Volsko Christina, Dudman Jessica, Christie Emilie, Dunham Jordon, Fox Robert J, Trapp Bruce D
Department of Neurosciences, Lerner Research Institute, Cleveland Clinic.
Department of Neurosciences, Lerner Research Institute, Cleveland Clinic; Mellen Center for Treatment and Research in Multiple Sclerosis, Neurological Institute, Cleveland Clinic.
J Vis Exp. 2019 Jul 19(149). doi: 10.3791/59511.
We describe a rapid tissue donation program for individuals with multiple sclerosis (MS) that requires scientists and technicians to be on-call 24/7, 365 days a year. Participants consent to donate their brain and spinal cord. Most patients were followed by neurologists at the Cleveland Clinic Mellen Center for MS Treatment and Research. Their clinical courses and neurological disabilities are well-characterized. Soon after death, the body is transported to the MS Imaging Center, where the brain is scanned in situ by 3 T magnetic resonance imaging (MRI). The body is then transferred to the autopsy room, where the brain and spinal cord are removed. The brain is divided into two hemispheres. One hemisphere is immediately placed in a slicing box and alternate 1 cm-thick slices are either fixed in 4% paraformaldehyde for two days or rapidly frozen in dry ice and 2-methylbutane. The short-fixed brain slices are stored in a cryopreservation solution and used for histological analyses and immunocytochemical detection of sensitive antigens. Frozen slices are stored at -80 °C and used for molecular, immunocytochemical, and in situ hybridization/RNA scope studies. The other hemisphere is placed in 4% paraformaldehyde for several months, placed in the slicing box, re-scanned in the 3 T magnetic resonance (MR) scanner and sliced into centimeter-thick slices. Postmortem in situ MR images (MRIs) are co-registered with 1 cm-thick brain slices to facilitate MRI-pathology correlations. All brain slices are photographed and brain white-matter lesions are identified. The spinal cord is cut into 2 cm segments. Alternate segments are fixed in 4% paraformaldehyde or rapidly frozen. The rapid procurement of postmortem MS tissues allows pathological and molecular analyses of MS brains and spinal cords and pathological correlations of brain MRI abnormalities. The quality of these rapidly-processed postmortem tissues (usually within 6 h of death) is of great value to MS research and has resulted in many high-impact discoveries.
我们描述了一项针对多发性硬化症（MS）患者的快速组织捐赠计划，该计划要求科学家和技术人员一年365天、每天24小时随叫随到。参与者同意捐赠他们的大脑和脊髓。大多数患者由克利夫兰诊所梅伦MS治疗与研究中心的神经科医生进行跟踪。他们的临床病程和神经功能障碍特征明确。患者死亡后不久，尸体被运至MS成像中心，在那里通过3T磁共振成像（MRI）对大脑进行原位扫描。然后尸体被转移到解剖室，在那里取出大脑和脊髓。大脑被分成两个半球。一个半球立即放入切片盒中，每隔1厘米厚的切片要么在4%多聚甲醛中固定两天，要么在干冰和2-甲基丁烷中快速冷冻。短期固定的脑切片保存在冷冻保存溶液中，用于组织学分析和敏感抗原的免疫细胞化学检测。冷冻切片保存在-80°C，用于分子、免疫细胞化学以及原位杂交/RNAscope研究。另一个半球置于4%多聚甲醛中几个月，然后放入切片盒，在3T磁共振（MR）扫描仪中重新扫描，并切成1厘米厚的切片。死后原位MR图像与1厘米厚的脑切片进行配准，以促进MRI与病理学的相关性研究。所有脑切片都进行拍照，并识别脑白质病变。脊髓被切成2厘米长的节段。每隔一段节段固定在4%多聚甲醛中或快速冷冻。快速获取死后的MS组织可对MS大脑和脊髓进行病理和分子分析，并对脑MRI异常进行病理相关性研究。这些快速处理的死后组织（通常在死亡后6小时内）的质量对MS研究具有重要价值，并促成了许多重大发现。
