Encyclopedia of Experiments: Biology
A subscription to JoVE is required to view this content. Sign in or start your free trial.
The JoVE video player is compatible with HTML5 and Adobe Flash. Older browsers that do not support HTML5 and the H.264 video codec will still use a Flash-based video player. We recommend downloading the newest version of Flash here, but we support all versions 10 and above.
If that doesn't help, please let us know.
Inbedding van volwassen drosophilahoofden in paraffine: weefsel voorbereiden op histologie
Transcript
Please note that all translations are automatically generated.
Click here for the English version.
- Om vliegen te immobiliseren voor paraffine-inbedding, duwt u de verdoofde Drosophila voorzichtig door de opening van een vliegenkraag, zodat hun hoofden gelijkmatig op het apparaat worden geplaatst terwijl ze nog steeds aan de rest van het lichaam zijn bevestigd. Voor weefselfixatie plaatst u de halsbanden in de oplossing van Carnoy. Carnoy's is een denaturerend fixatiemiddel dat bestaat uit ethanol, chloroform en azijnzuur.
Om het monster voor te bereiden op paraffine-infiltratie, droogt u het weefsel uit door de kraag over te brengen door gesorteerde concentraties ethanol. Breng vervolgens de kraag over naar methylbenzoaat, een organisch oplosmiddel. Plaats het vervolgens in een methylbenozoaat en gesmolten paraffineoplossing terwijl het wordt verwarmd om ervoor te zorgen dat de paraffine gesmolten blijft. Ga verder met een uitgebreide reeks wasstappen met alleen voorverwarmde paraffine om het methylbenzoaat te verwijderen.
Plaats vervolgens snel de kraag in een mal en giet gesmolten paraffine om deze volledig te bedekken. Zodra de paraffine is uitgehard, verwijdert u het blok uit de mal en gebruikt u een scheermesje om overtollige paraffine te trimmen. Breek de kraag af en laat de rij ingebedde hoofden in het paraffineblok achter. In het voorbeeldprotocol zien we de procedure gedemonstreerd voor de analyse van neurodegeneratieve fenotypes.
- Gebruik eerst een tang om de onderwerpvliegen bij de nek in de halsbanden te rijgen. Lijn alle hoofden in dezelfde richting uit en zorg ervoor dat er geen schade aan het hoofd of de ogen optreedt. Neem oogloze sinus oculisvliegen op willekeurige bekende posities op, zodat de volgorde van de vliegen gemakkelijk in de secties kan worden geïdentificeerd. Bovendien, als de vliegbelasting lichte of witte ogen heeft, draad roodogige vliegen met tussenpozen langs de strip om ervoor te zorgen dat er voldoende pigment aanwezig is om de dia te bevlekken.
Noteer de volgorde van de vliegen samen met het halsbandnummer als er meer dan één wordt gebruikt. Zodra een halsband klaar is, plaatst u deze gedurende 3,5 tot 4 uur in de oplossing van Carnoy. Voer een reeks ethanol-, methylbenzoaat- en paraffinewassingen uit om het vliegweefsel voor te bereiden op inbedding. Plaats vervolgens de halsbanden één per sectie in een rubberen ijsblokjesbak.
Giet voorzichtig gesmolten paraffine over de kragen en vermijd luchtbellen en laat het 's nachts uitharden. Verwijder de paraffineblokken met de kragen uit de lade. Scheid het paraffineblok van de kraag met een scheermesje en breek de kraag voorzichtig af. De lichamen blijven in de kraag terwijl de hoofden in het paraffineblok blijven.
Tags
Lege waarde probleemRelated Videos
PLAYLIST
-
Education • BiochemistryPhotometric Protein Determination
-
Education • Basic Methods in Cellular and Molecular BiologySeparating Protein with SDS-PAGE
-
Education • Basic Methods in Cellular and Molecular BiologyThe Western Blot
-
Education • Basic Methods in Cellular and Molecular BiologyThe ELISA Method
-
Education • ImmunologyImmunoprecipitation-Based Techniques: Purification of Endogenous Proteins Using Agarose Beads
-
Education • Basic Methods in Cellular and Molecular BiologyPCR: The Polymerase Chain Reaction
-
Education • Basic Methods in Cellular and Molecular BiologyDNA Gel Electrophoresis
-
Education • Basic Methods in Cellular and Molecular BiologyBacterial Transformation: The Heat Shock Method
-
Education • Basic Methods in Cellular and Molecular BiologyPlasmid Purification
-
Education • Cell BiologyLive Cell Imaging of Mitosis
-
Education • Cell BiologyThe Transwell Migration Assay
-
Education • Cell BiologyCell-surface Biotinylation Assay
-
Education • Cell BiologyFM Dyes in Vesicle Recycling
-
Education • Cell BiologyThe ATP Bioluminescence Assay
-
Education • BiochemistryMetabolic Labeling
-
Education • Cell BiologyThe TUNEL Assay
-
Education • Cell BiologyAnnexin V and Propidium Iodide Labeling
-
Research • BiologyViability Assays for Cells in Culture
-
Education • ImmunologyAssay for Cell Death: Chromium Release Assay of Cytotoxic Ability
-
Education • Basic Methods in Cellular and Molecular BiologyPassaging Cells
-
Education • Basic Methods in Cellular and Molecular BiologyUsing a Hemacytometer to Count Cells
-
Research • BiologyDetermination of Mammalian Cell Counts, Cell Size and Cell Health Using the Moxi Z Mini Automated Cell Counter
-
Education • Basic Methods in Cellular and Molecular BiologyAn Introduction to Transfection
-
Education • NeurosciencePrimary Neuronal Cultures
-
Education • NeuroscienceNeuronal Transfection Methods
-
Research • Developmental BiologyCulture and Transfection of Zebrafish Primary Cells
-
Research • Immunology and InfectionHighly Efficient Transfection of Primary Macrophages with In Vitro Transcribed mRNA
-
Research • BiologyHigh Yield Expression of Recombinant Human Proteins with the Transient Transfection of HEK293 Cells in Suspension
-
Education • Developmental BiologyEmbryonic Stem Cell Culture and Differentiation
-
Education • Developmental BiologyInduced Pluripotency
-
Education • Biology IIBasic Mouse Care and Maintenance
-
Education • Lab Animal ResearchRodent Handling and Restraint Techniques
-
Education • Lab Animal ResearchCompound Administration I
-
Education • Lab Animal ResearchCompound Administration II
-
Education • Lab Animal ResearchCompound Administration III
-
Education • Lab Animal ResearchCompound Administration IV
-
Research • BiologyManual Restraint and Common Compound Administration Routes in Mice and Rats
-
Education • Biology IIIntroducing Experimental Agents into the Mouse
-
Research • BehaviorSystematic Assessment of Well-Being in Mice for Procedures Using General Anesthesia
-
Education • Lab Animal ResearchBlood Withdrawal I
-
Education • Lab Animal ResearchBlood Withdrawal II
-
Education • Lab Animal ResearchAnesthesia Induction and Maintenance
-
Education • Lab Animal ResearchConsiderations for Rodent Surgery
-
Research • NeuroscienceWhole Animal Perfusion Fixation for Rodents
-
Education • Lab Animal ResearchDiagnostic Necropsy and Tissue Harvest
-
Education • General Laboratory TechniquesIntroduction to Light Microscopy
-
Research • BiologyPhase Contrast and Differential Interference Contrast (DIC) Microscopy
-
Education • General Laboratory TechniquesIntroduction to Fluorescence Microscopy
-
Education • Analytical ChemistryScanning Electron Microscopy (SEM)
-
Research • BiochemistryImaging of Extracellular Vesicles by Atomic Force Microscopy
-
Education • ImmunologyConfocal Fluorescence Microscopy: A Technique to Determine the Localization of Proteins in Mouse Fibroblasts
-
Research • BioengineeringVideo-rate Scanning Confocal Microscopy and Microendoscopy
-
Research • BiologyThe CryoAPEX Method for Electron Microscopy Analysis of Membrane Protein Localization Within Ultrastructurally-Preserved Cells
-
Education • General Laboratory TechniquesHistological Sample Preparation for Light Microscopy
-
Education • NeuroscienceHistological Staining of Neural Tissue
-
Research • Encyclopedia of Experiments: BiologyNile Red Staining of C. elegans: A Method to Visualize Lipid Droplets in Fixed Animals
-
Research • Encyclopedia of Experiments: BiologyEthanol Fixation and DAPI Staining: A Method to Visualize DNA in C. elegans
-
Research • NeurosciencePreparation of Non-human Primate Brain Tissue for Pre-embedding Immunohistochemistry and Electron Microscopy
-
Research • Encyclopedia of Experiments: BiologyEmbedding Adult Drosophila Heads in Paraffin: Preparing Tissue for Histology
-
Research • BiologyCryosectioning of Contiguous Regions of a Single Mouse Skeletal Muscle for Gene Expression and Histological Analyses
-
Research • MedicineSampling Strategies and Processing of Biobank Tissue Samples from Porcine Biomedical Models
-
Education • ImmunologyImmunohistochemistry and Immunocytochemistry: Tissue Imaging via Light Microscopy
-
Education • ImmunologyImmunofluorescence Microscopy: Immunofluorescence Staining of Paraffin-Embedded Tissue Sections
-
Research • NeuroscienceImmunostaining of Biocytin-filled and Processed Sections for Neurochemical Markers
-
Education • Developmental BiologyWhole-Mount In Situ Hybridization
-
Education • Core: ChemistryUncertainty in Measurement: Accuracy and Precision
-
Education • Analytical ChemistryInternal Standards
-
Education • Analytical ChemistryCalibration Curves
-
Education • Analytical ChemistryMethod of Standard Addition
-
Education • Analytical ChemistrySample Preparation for Analytical Characterization
-
Education • Analytical ChemistryIntroduction to Mass Spectrometry
-
Education • BiochemistryTandem Mass Spectrometry
-
Research • BioengineeringSample Preparation Strategies for Mass Spectrometry Imaging of 3D Cell Culture Models
-
Research • GeneticsMass Spectrometry-Based Proteomics Analyses Using the OpenProt Database to Unveil Novel Proteins Translated from Non-Canonical Open Reading Frames
-
Education • Materials EngineeringX-ray Diffraction
-
Education • Analytical ChemistryRaman Spectroscopy for Chemical Analysis
-
Education • Analytical ChemistryUltraviolet-Visible (UV-Vis) Spectroscopy
-
Research • EngineeringQuasi-light Storage for Optical Data Packets
-
Education • Analytical ChemistryHigh-Performance Liquid Chromatography (HPLC)
-
Education • Analytical ChemistryGas Chromatography (GC) with Flame-Ionization Detection
-
Education • Biology IIMouse Genotyping
-
Education • GeneticsSNP Genotyping
-
Research • BiologyRapid Genotyping of Mouse Tissue Using Sigma's Extract-N-Amp Tissue PCR Kit
-
Research • BiologyInfinium Assay for Large-scale SNP Genotyping Applications
-
Research • BiologyA PCR-based Genotyping Method to Distinguish Between Wild-type and Ornamental Varieties of Imperata cylindrica
-
Research • Developmental BiologyGenotyping and Quantification of In Situ Hybridization Staining in Zebrafish
-
Research • MedicineCollection and Extraction of Saliva DNA for Next Generation Sequencing
-
Research • BiologyAmplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites
-
Research • BiologyNext-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
-
Research • BiologyTargeted DNA Methylation Analysis by Next-generation Sequencing
-
Education • Core: PsychologyMeasures of Central Tendency
-
Education • Core: PsychologyVariation: Normal Distribution, Range, and Standard Deviation
-
Education • Core: PsychologyRegression Toward the Mean
-
Education • Core: PsychologyStatistical Significance
-
Research • NeuroscienceStatistical Modelling of Cortical Connectivity Using Non-invasive Electroencephalograms
-
Research • BiochemistryFully Autonomous Characterization and Data Collection from Crystals of Biological Macromolecules
-
Research • BehaviorAutomated, Quantitative Cognitive/Behavioral Screening of Mice: For Genetics, Pharmacology, Animal Cognition and Undergraduate Instruction
-
Research • Cancer ResearchIntegration of Bioinformatics Approaches and Experimental Validations to Understand the Role of Notch Signaling in Ovarian Cancer
-
Research • BiologyVideo Bioinformatics Analysis of Human Embryonic Stem Cell Colony Growth
-
Research • GeneticsTargeted Next-generation Sequencing and Bioinformatics Pipeline to Evaluate Genetic Determinants of Constitutional Disease
-
Research • GeneticsRNA Next-Generation Sequencing and a Bioinformatics Pipeline to Identify Expressed LINE-1s at the Locus-Specific Level
-
Research • GeneticsA Bioinformatics Pipeline to Accurately and Efficiently Analyze the MicroRNA Transcriptomes in Plants
-
Research • BiologyA Bioinformatics Pipeline for Investigating Molecular Evolution and Gene Expression using RNA-seq