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Histology Lab 2013
| Question | Answer |
|---|---|
| 4 tissue types recognized in histology | Epithelium, Connective tissue, Muscle, Nervous |
| Use of formaldehyde | Main fixative of histology. Reacts with proteins to preserve tissues. No reaction to lipids. |
| Use of Glutaldehyde | Has less penetrance but good for the thin slices that are used for electron microscope |
| Use of frozen sectioning | Good for rapid prognosis when needing a quick diagnosis while in surgery. Also good for staining with antibodies. Quick and dirty procedure |
| What must take place in order to fix with formaldehyde? | Dehydration of sample using graded alcohol series (to prevent rapid shrinking). Done because parafin doesn't work well with water. |
| Hemotoxylin | Major stain of histology. Stains acidic structures (reacts primarily with DNA) |
| Eosin | Major stain of histology. Stains basic structures. Generally stains proteins. |
| Steps of Kohler illumination | 1.Focus the objective of specimen 2.Close the field diaphragm 3.Center the field diaphragm by adjusting the position of the condensor 4.Focus the image of the field diaphram by moving condensor up and down 5.Open field dia. and adjust aperture dia. |
| 3 main steps of tissue preparation | 1. fixation 2. embedding in paraffin to permit sectioning 3. staining |
| Main fixative for electron microscopy | Heavy metal (Osmium tetroxide) |
| Parafin | Embedding medium. Allows for easy slicing of specimen. |
| Autoradiography | Radioactively tagged precursors of a molecule are incorporated by cells and tissues before fixation. Makes use of a photographic emulsion placed over a tissue section to localize radioactive materials within tissues |
| Ionic charges as related to H & E | -Eosin is an acidic dye and carries a net negative charge (acidophilic component binding) -Hematoxylin is (more or less) a basic dye and carries a net positive charge (basophilic component binding) |
| Dyes of the Mallory Staining technique and the components they are used on | aniline blue (for collagen), acid fuchsin (for ordinary cytoplasm and nuclei), and orange G (for red blood cells) |
| Substances within cells and the extracellular matrix which display basophilia | -heterochromatin/ nucleoli -cytoplasmic components -extracellular materials |
| Substances within cells and the extracellular matrix which display acidophilia | -cytoplasmic filaments -intracellular membranous components -extracellular fibers |
| PAS Reaction | Periodic Acid-Schiff reaction: Stains carbohydrates and carbohydrate-rich macromolecules. It is used to stain glycogen in cells, mucus in cells/tissues, the basement membrane which underlies epithelia, and reticular fibers in connective tissue |
| Feulgen Reaction | Used to stain DNA |
| Method to quantify amount of DNA in cell nucleus | PAS. Reaction of Schiff reagent with DNA is stoichiometric; proportional to amount of DNA. WONT work with RNA (no deoxyribose) |
| Direct Immunofluorescence | Somewhat obsolete, single-step procedure that involves a single labeled antibody. Visualization of structures is not ideal due to low intensity of the signal emission |
| Indirect Immunofluorescence | Labeling in which the first antibody is unconjugated, but is labeled by a second, antiimunoglobulin antibody, which is conjugated to a fluorochrome or enzyme |
| Advantages to Indirect Immunofluorescence | -It considerably enhances the fluorescence signal emission from the tissue - A single secondary antibody can be used to localize the tissue-specific binding of several different primary antibodies |
| Resolving Power | The ability of a microscope lens or optical system to produce separate images of closely positioned objects |
| With wavelength of 540, under ideal conditions, what is the max resolving power of a bright-field microscope? | .2μm |
| Factors which help to determine resolution | Optical System, wavelength of light source, specimen thickness, quality of fixation, and staining intensity |
| Relation between ocular lens and resolution | There is no true relationship. Ocular (eyepiece) magnifies the image produced by the objective lens, but it CANNOT increase resolution |
| Artifact | Error in the preparation process when forming a histological slide |
| Phase contrast microscope | Translates differences in refractive indexes into differences in light intensity. Ideal for unstained cells and tissues; particularly living cells |
| Dark-field microscopy | Light scattering at cell boundaries; useful in examining autoradiographs, in which the developed silver grains appear white in a dark background |
| Fluorescence Microscope | Uses set of filters to see natural or introduced fluorescent molecules |
| Confocal Microscopy | Fluorescence microscopy using a computer-guided laser beam which scans the specimen in 3 dimensions (and has 2-dimensional capability as well). May be used without specimen fixation |
| UV microscope | Uses UV light to detect nucleic acids. Does not use a objective lens because UV light is not visible, so instead is only viewable photographically |
| Polarizing microscope | Analyzes angle changes in polarized light to determine the degree with which a structure affects the light (good for striated muscle) |
| TEM | Transmission Electron Microscopy. Similar to light except higher resolution due to usage of electrons. Also uses magnetic lenses. Final image appears on fluorescent screen or photographic plate |
| Specimen prep for TEM | 1. Fixation (cross-linking) using glutaraldehyde. 2.Buffering 3. Embedding in plastic (epoxy resin) 4. Cut to thin slices using diamond or glass knife 5. Staining with heavy metal ion (osmium tetroxide often times) Freeze fracture method used somet |
| SEM | Scanning Electron Microscopy. Provides for 3-dimensional, "TV like" image. Uses beam of electrons to scan surfaces of specimen which has been coated in gold. Electron detector detects deflected electrons |
| Specimen prep for SEM | 1. Fixation 2. Dehydration 3. Coating in evaporated gold-carbon film 4. Mounting on aluminum stub 5.Placement in SEM specimen chamber |
| Main disadvantage of TEM | Small penetrating power of electrons means samples must be observed in a high vacuum and must be extremely thinly sliced |
| Neuroglial Cells of the CNS | (1)astrocytes (2) microglial cells (3)oligodendrocytes (4)ependymal cells |
| Neuroglial Cells of the PNS | (1)Schwann cells (2)satellite cells |
| Relationship between resolving power and Numerical Aperture # and R (in nm)? | Direct. The larger the NA the higher the resolving power and the smaller R in nm |
| Nissl substance | Dense concentrations of RER and polyribosomes |
| Perineurium | Connective tissues which directly surrounds bundles of nerve fibers |
| Epineurium | Connective tissue which surrounds several bundles of nerve fibers |
| Nerve vs Neuron | Nerve= Bundle of axons Neuron= Nerve cell (with 1 or 2 axons) |
| Components of the PNS | Sensory and Motor. Motor is broken into Sensory and Autonomic |
| Staining which makes myelin appear Black | Osmium Tetroxide |
| Endonerium | Connective tissue between individual axons |
| Neuropil | Network of axons, dendrites, and neuroglial processes |
| Blood-Brain barrier | Endothelial cells lining the continuous capillaries coursing through the CNS form tight junctions that prevent the free flow of blood-bourne substances into the neural tissue |
| CSF | Cerebral Spinal Fluid. Produced by Choroid Plexus |
| How many layers does the cerebrum have? | 6 |
| What does the cerebellum do? | Coordinates voluntary movements & muscle function in the maintenance of normal posture. |
| Layers of Cerebellum | Molecular, purkinje, Granular (grey), white |
| Folia | One branch of cerebellum |
| 2 forms of Ganglia | -Sensory -Autonomic |
| Ganglia | Groups of neuron cell bodies which occur outside of the CNS |
| Cells associated with sensory ganglia | pseudounipolar, bipolar neurons |
| Cells associated with autonomic ganglia | sympathetic & parasympathetic multipolar neurons |
| Autonomic ganglia subclasses | Sympathetic, Parasympathetic and Enteric Ganglia |
| What are the oval structures just under the submucosa? | Submucosal plexus |
| Type of connective tissue in a tendon | Dense irregular |
| What cells do chondroblasts and Osteoblasts originate from | Mesenchymal cells |
| Appositional vs Interstitial growth | Appositional- the process that forms new cartilage at the surface of an existing cartilage Interstitial growth- the process that forms new cartilage within an existing cartilage mass |
| Location of fibrocartilage | Between vertebral disks |
| What cells do osteoclasts originate from? | monocytes |
| How do individual osteocytes receive blood supply | canaliculi |
| When doing a transverse crossection of bone, what are the horizontal openings that are visible by the haversion canals | Volksmann's canals |
| Circular component which surrounds haversian canal | Osteon |
| Zones involved in endochondral bone growth | (1)Reserve cartilage (2)Zone of proliferation (3)zone of maturation (4) Zone of hypertrophy and calcification (5)Cartilage degeneration (6)osteogenesis |
| What is the name for the bits of cartilage in newly forming bone | cartilage spicules |
| What do fibroblasts make? | Extracellular matrix and fibers |
| Proteoglycan | Proteins which are heavily gylcosylated. Found in connective tissue. Major component of extracellular matrix. They form large complexes to other proteoglycans, to hyaluronan and to fibrous proteins. Effect signaling/stability. |
| What is responsible for giving tissues their elasticity? | Elastin. A highly hydrophobic protein which is unusually rich in proline and glycine. |
| Dominant extracellular matrix of arteries | Elastin |
| What allows the elastic network to stretch and recoil like a rubber band? | Random coil formation of parts of the elastin polypeptide chain cross-linked into the elastic fiber network |
| Structure of Fibronectin | It is a dimer composed of two very large subunits joined by disulfide bonds at one end. Each subunit is folded into a series of functionally distinct domains seperated by regions of flexible polypeptide chain. |
| The structure of laminin | Laminin is a multidomain glycoprotein composed of three polypeptides (alpha, beta, and gamma)that are disulfide-bonded into an asymmetric crosslike structure |
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