辣根过氧化物酶标记与样品后染色联用在微观脑联结组学中的应用

Application of Horseradish Peroxidase Labeling Along With Post-Staining Improves Data Quality in Micro-Brain Connectomics

三维电子显微成像(volume electron microscopy)是微观尺度脑联结组学(micro-brain connectomics)研究的主要研究手段.自动卷带式连续超薄切片-收片系统(automated tape-collecting ultramicrotome,ATUM)及扫描电镜序列成像系统的联合应用,是三维电镜成像中一种重要的技术手段.该方法利用卷带式收集超薄切片,形成完整的样品切片库,进而在后续成像中形成样品三维电镜成像数据库.根据ATUM系统镜外切片-收片、保留样本切片库等特点,对相应的生物样品制备方法进行改进,包括树脂配方及树脂渗透方式改良,神经元细胞膜辣根过氧化物酶(horseradish peroxidase,HRP)稀疏化标记与样本切片库后染色联用——从而很大程度上减少样品切片的褶皱率,并使得在整体高衬度的样品图像库中高效地识别神经元身份成为可能,最终获得符合脑联结组学所需的高质量成像数据.

For micro-brain connectomic research,volume electron microscopy is a main method in which it plays an important role for the application of automated tape-collecting ultramicrotome(ATUM)system and array imaging with scanning electron microscopy together.ATUM system enables collecting a large amount of serial sections to be imaged with SEM to form a huge dataset.With regard to the nature of non-vacuum sectioning and sample reserving,we modified and improved the sample preparation process including resin recipe,HRP labelling and post-staining method in order to reduce folding rate of sample sections and make it possible for identifying certain types of neurons in high-contrast imaging dataset,thus obtaining dataset of high quality for connectomic research.