Formalin fixed Paraffin Embedded (FFPE) Tissue preparation for digital pathology quantification using QuPath
By Emma Sherrell onQuPath is an open-source digital pathology analysis software. Here, it is used to quantify various misfolded proteins across anatomical regions. This protocol is appropriate for both human and mouse tissue, and will outline the methods of tissue preparation, digitisation, segmentation, and importing images into QuPath.
Habituation and dishabituation (olfaction test)
By Emma Sherrell onHabituation and dishabituation (olfaction test) for mice.
Targeted detection of SNCA CNVs in SOX10+ nuclei from oligodendrocytes containing alpha-synuclein inclusions isolated from human post-mortem brain
By savannah onThe protocol is optimized to detect SOX10, a nuclear oligodendrocyte marker, and alpha-synuclein inclusions, which are frequently retained at the perinucleus in MSA (the so-called Papp-Lantos inclusions). This protocol also describes its use in affected PD and MSA brain regions such as the putamen, substantia nigra (SN), and cerebellum.
Section 3: Libraries quality control (QC)
By savannah onThis protocol details quality control of libraries and should be performed after Section 2: NGS library preparation for sequencing.
Section 2: NGS library preparation for sequencing
By savannah onThis protocol details NGS library preparation for sequencing and should be performed after Section 1: Enzymatic DNA Fragmentation (Manually).
Section 1: Enzymatic DNA Fragmentation (Manually)
By savannah onThis protocol details manual enzymatic DNA Fragmentation prior to Section 2: NGS library preparation for sequencing.
Manual isolation of nuclei from human brain using CellRaft device and single nucleus Whole Genome Amplification
By savannah onProtocol for manual nuclear isolation from human brain tissue using Cell Raft device for single cell Whole Genome Amplification Protocol for manual nuclear isolation from human brain tissue using Cell Raft device for single cell Whole Genome Amplification
Single-cell Whole Genome Amplification (scWGA) of human post-mortem brain samples isolated by Laser Capture Microdissection (LCM)
By savannah onThis protocol uses Laser Capture Microdissection (LCM) technology on human post-mortem brain tissue slides with rapid Giemsa staining to isolate single nuclei for Single-cell Whole Genome Amplification (scWGA) in order to do low coverage (<1x) single-cell whole genome sequencing to detect mega-base somatic Copy Number Variations (CNVs).
Gavage – food supplementation for stalled weight gain in mice
By Emma Sherrell onFeeding protocol for Ndufs2fl/fl mice,
Preparation of tissue for Ribo-Tag/RNAseq analysis
By Emma Sherrell onThis protocols describes the RiboTag virus vectors injected into mouse brain, tissue dissection, immunoprecipitation and RNA extraction for RNAseq.
Sleep-wake recording in Dan Lab
By Emma Sherrell onSurgery and setups for recording 24-hour natural sleep/wake cycles in mice.
Standard operating procedure (SOP) for combined ICA and systemic administration of MPTP: The overlesioned (Bilateral Asymmetric) non-human primate model
By Emma Sherrell onThis SOP describes the procedures for the combined internal carotid artery (ICA) and systemic (intravenous or intramuscular) administration of MPTP in non-human primates. This model produces an overlesioned or bilateral asymmetric model, induced by unilateral ICA plus systemic administration. This protocol has been adapted from Bankiewicz et al. (1999).
Electrophysiological recording from a chronic chamber-implanted non-human primate
By Emma Sherrell onThis protocol describes the preparation and insertion of extracellular recording electrodes into intracranial targets of a non-human primate implanted with a chronic recording chamber. A piercing guide cannula is used to penetrate the dura and to guide the electrode. Practices related to targeting, electrode cleaning, and electrode movement and settling relative to brain tissue are discussed.
Essential surgeries for the electrophysiological recording from a behaving non-human primate brain 2 (Craniotomy)
By Emma Sherrell onOutlined are the steps to complete a craniotomy in an existing recording chamber placed on a cranial implant. This procedure can be done concurrently with the implant surgery (not recommended), or can take place upon recovery from the implant surgery.
Essential surgeries for the electrophysiological recording from a behaving non-human primate brain 1 (Head fixation post and recording chamber implantation)
By Emma Sherrell onThe performing surgical procedures under strictly sterile conditions is the key to the successful progress of the project.
Maintenance of the recording chamber for non-human primate (Chamber cleaning and dura scraping)
By Emma Sherrell onOnce a craniotomy has been done in the chamber, the inside of the chamber must be cleaned regularly (2-3 times per week) to prevent infection. This protocol explains two procedures to maintain the recording chamber in a healthy condition:(1) chamber cleaning and (2) dura scraping.
DAB antibody (IHC) staining protocol
By Emma Sherrell onThis is the basic protocol for antibody staining of formalin fixed paraffin embedded (FFPE) tissue. This is the basic protocol for antibody staining of formalin fixed paraffin embedded (FFPE) tissue.