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Questions and Answers
Which staining method is used to distinguish between different types of extracellular tissue components?
What is the primary purpose of the Periodic acid-Schiff (PAS) reaction in histology?
How does Sudan black staining differ from other histological staining techniques?
What is the primary purpose of metal impregnation techniques in histology?
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Which staining method uses a three-color system to emphasize support fibers in connective tissue?
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What is the primary function of hematoxylin and eosin (H&E) staining in histology?
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What is the primary purpose of the pinhole aperture in a confocal microscope?
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Which of the following is a characteristic of basic dyes used in histology?
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Which of the following cell components would be stained by an acidic dye in histology?
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How does the use of a small point of high-intensity light, often from a laser, in confocal microscopy improve resolution?
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What is the primary difference between the staining properties of basic and acidic dyes in histology?
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How does the use of a confocal microscope improve the localization of specimen components compared to traditional fluorescence microscopy?
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Which type of microscopy allows for the visualization of thin optical sections within a thick specimen?
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Which staining technique is commonly used in fluorescence microscopy to visualize specific cellular components?
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Which type of dyes are generally used to stain acidic components in cells and tissues?
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Which cellular component is typically stained by the basic dye hematoxylin in histological preparations?
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Which staining technique is commonly used to visualize glycogen and mucopolysaccharides in tissues?
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What is the primary purpose of the fixation step in tissue preparation for light microscopy?
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Study Notes
Staining and Stains
- Trichrome stains allow for greater distinctions among various extracellular tissue components, such as Masson trichrome.
- The periodic acid-Schiff (PAS) reaction utilizes the hexose rings of polysaccharides and other carbohydrate-rich tissue structures, staining them purple or magenta.
- Sudan black is a lipid-soluble dye that stains lipids, avoiding processing steps that remove lipids.
- Metal impregnation using solutions of silver salts can visualize specific ECM fibers and cellular elements in nervous tissue.
Commonly Used Histological Stains
- Hematoxylin and eosin (H&E): nucleus/blue, cytoplasm/pink.
- Van Gieson method: collagen/pink, muscle/yellow.
- Trichrome method: three-color system to emphasize support fibers: connective tissue/blue, cytoplasm/pink, nuclei/dark brown.
Tissue Types
- Epithelium
- Connective tissue
- Muscular tissue
- Nervous tissue
Electron Microscopy
- Confocal microscopy achieves high resolution and sharp focus using a small point of high-intensity light and a plate with a pinhole aperture.
- The point light source, focal point of the lens, and detector's pinpoint aperture are optically conjugated.
Staining
- Most cells and extracellular material are completely colorless.
- Dyes form electrostatic (salt) linkages with ionizable radicals of macromolecules in tissues.
- Basic dyes (e.g., toluidine blue, alcian blue, methylene blue) stain basophilic components (e.g., nucleic acids).
- Acid dyes (e.g., eosin, orange G, acid fuchsin) stain acidophilic components (e.g., proteins).
Histology
- Histology is the study of the tissues of the body and how these tissues are arranged to constitute organs.
- Tissue is composed of cells and ECM (extracellular matrix).
- Levels of organization: tissue → organ → organism.
Tissue Processing for Histology
- Fixation: small pieces of tissue are placed in solutions that cross-link proteins and inactivate degradative enzymes.
- Dehydration: the tissue is transferred through a series of increasingly concentrated alcohol solutions, removing all water.
- Clearing: alcohol is removed in organic solvents in which both alcohol and paraffin are miscible.
- Infiltration: the tissue is placed in melted paraffin until it becomes completely infiltrated.
- Embedding: the paraffin-infiltrated tissue is placed in a small mold with melted paraffin and allowed to harden.
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Description
Learn about the principles of confocal microscopy, which achieves high resolution and sharp focus by using a small point of high-intensity light and a pinhole aperture. Explore how optical conjugation in the focal plane improves resolution of the object being observed.
