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换一换DAB Immunohistochemistry (IHC) Staining for Stereological Analysis
Nicolas Giguère, louis-eric.trudeau Trudeau
DAB (3,3'-diaminobenzidine) is oxidized in the presence of peroxidase and hydrogen peroxide resulting in the deposition of a brown, alcohol-insoluble precipitate at the site of enzymatic activity.DAB (3, 3'-diaminobenzidine) produces a dark brown reaction product and can be used for immunohistochemical and applications. This protocol details the DAB immunohistochemistry staining for stereological analysis of 40 µM slices cut with cryostat and stored in antifreeze.
AI 解读Human Neuronal Nucleus Isolation for Single-Nucleus Transcriptomic Profiling (10x Genomics)
Satoshi Ishishita, Allan-Hermann Pool
Protocol generating suspensions of neuronal nuclei (NeuN+) from human central nervous system tissue for single-nucleus transcriptomics.
AI 解读Analysis of nanomaterial biocoronas in biological and environmental surroundings
Peng Zhang, Mingjing Cao, Andrew J. Chetwynd, Klaus Faserl, Fazel Abdolahpur Monikh, Wei Zhang, Rawi Ramautar, Laura-Jayne A. Ellis, Hossein Hayat Davoudi, Katie Reilly, Rong Cai, Korin E. Wheeler, Diego Stéfani Teodoro Martinez, Zhiling Guo, Chunying Chen, Iseult Lynch
A biomolecular coating, or biocorona, forms on the surface of engineered nanomaterials (ENMs) immediately as they enter biological or environmental systems, defining their biological and environmental identity and influencing their fate and performance. This biomolecular layer includes proteins (the protein corona) and other biomolecules, such as nucleic acids and metabolites. To ensure a meaningful and reproducible analysis of the ENMs-associated biocorona, it is essential to streamline procedures for its preparation, separation, identification and characterization, so that studies in different labs can be easily compared, and the information collected can be used to predict the composition, dynamics and properties of biocoronas acquired by other ENMs. Most studies focus on the protein corona as proteins are easier to monitor and characterize than other biomolecules and play crucial roles in receptor engagement and signaling; however, metabolites play equally critical roles in signaling. Here we describe how to reproducibly prepare and characterize biomolecule-coated ENMs, noting especially the steps that need optimization for different types of ENMs. The structure and composition of the biocoronas are characterized using general methods (transmission electron microscopy, dynamic light scattering, capillary electrophoresis–mass spectrometry and liquid chromatography–mass spectrometry) as well as advanced techniques, such as transmission electron cryomicroscopy, synchrotron-based X-ray absorption near edge structure and circular dichroism. We also discuss how to use molecular dynamic simulation to study and predict the interaction between ENMs and biomolecules and the resulting biocorona composition. The application of this protocol can provide mechanistic insights into the formation, composition and evolution of the ENM biocorona, ultimately facilitating the biomedical and agricultural application of ENMs and a better understanding of their impact in the environment.
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