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Questions and Answers
Which microscopy technique creates a pseudo-3D effect by using polarized light?
Which microscopy technique creates a pseudo-3D effect by using polarized light?
Stains improve contrast in live cell imaging.
Stains improve contrast in live cell imaging.
False
What is the primary method used to enhance contrast in conventional bright field microscopy?
What is the primary method used to enhance contrast in conventional bright field microscopy?
Illuminating the specimen with white light
In differential interference contrast optics, the interference between two closely spaced beams generates __________ when the refractive index changes.
In differential interference contrast optics, the interference between two closely spaced beams generates __________ when the refractive index changes.
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Match each microscopy technique with its characteristic:
Match each microscopy technique with its characteristic:
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What thickness are typical sections when using a microtome for light microscopy?
What thickness are typical sections when using a microtome for light microscopy?
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Optical techniques can improve contrast for both fixed and live cells.
Optical techniques can improve contrast for both fixed and live cells.
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Name one advantage of using phase contrast microscopy over conventional bright field microscopy.
Name one advantage of using phase contrast microscopy over conventional bright field microscopy.
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What is the typical maximum resolution range for Electron Microscopy instruments?
What is the typical maximum resolution range for Electron Microscopy instruments?
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Freeze-fracture EM is used primarily to determine the external structure of a cell.
Freeze-fracture EM is used primarily to determine the external structure of a cell.
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What method is used to remove water from tissues before examining them with Conventional SEM?
What method is used to remove water from tissues before examining them with Conventional SEM?
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In Atomic Force Microscopy, the probe is a tiny moveable __________ that moves over the surface.
In Atomic Force Microscopy, the probe is a tiny moveable __________ that moves over the surface.
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Match the microscopy technique with its primary application:
Match the microscopy technique with its primary application:
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Which heavy metal is commonly used to coat surfaces in Conventional SEM to enhance conductivity?
Which heavy metal is commonly used to coat surfaces in Conventional SEM to enhance conductivity?
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Cryo EM uses high doses of electrons for imaging samples.
Cryo EM uses high doses of electrons for imaging samples.
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What type of samples are ideally examined with Atomic Force Microscopy?
What type of samples are ideally examined with Atomic Force Microscopy?
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What is the primary limitation of Scanning Electron Microscopy (SEM) compared to Transmission Electron Microscopy (TEM)?
What is the primary limitation of Scanning Electron Microscopy (SEM) compared to Transmission Electron Microscopy (TEM)?
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Transmission Electron Microscopy (TEM) can achieve magnification up to 1,000,000 times.
Transmission Electron Microscopy (TEM) can achieve magnification up to 1,000,000 times.
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What is the purpose of using immunogold labeling in electron microscopy?
What is the purpose of using immunogold labeling in electron microscopy?
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The technique that uses heavy metal salts to darken the background while contrasting with biological particles is called ___________ staining.
The technique that uses heavy metal salts to darken the background while contrasting with biological particles is called ___________ staining.
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Match the imaging methods with their characteristics:
Match the imaging methods with their characteristics:
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Which of the following macromolecules can be visualized using negative staining?
Which of the following macromolecules can be visualized using negative staining?
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Ultrastructure of a cell is not visible using Transmission Electron Microscopy (TEM).
Ultrastructure of a cell is not visible using Transmission Electron Microscopy (TEM).
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Name one fixative used in preparing tissue sections for Transmission Electron Microscopy.
Name one fixative used in preparing tissue sections for Transmission Electron Microscopy.
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Study Notes
Microscopy Techniques
- Microscopy is used to visualize structures at various scales, from atoms to cells and tissues.
- Light microscopy provides low-resolution images but is useful for viewing whole cells and organelles.
- Electron microscopy offers high resolution, visualizing subcellular structures and macromolecules.
- Different types of electron microscopy exist, including transmission electron microscopy (TEM) for visualizing internal structures and scanning electron microscopy (SEM) for visualizing surfaces.
- Atomic force microscopy (AFM) generates images of surfaces, biofilms, and molecules in solution.
- Newer super-resolution confocal microscopes overcome the 0.2 µm resolution limit of conventional light microscopes.
- Fluorescence microscopy uses fluorescent dyes or antibodies to visualize specific molecules within cells.
- Immunofluorescence staining enhances contrast using antibodies linked to fluorescent labels.
- Fluorescence in situ hybridization (FISH) localizes specific DNA or RNA sequences within cells.
- Proteins can be tagged in living cells using green fluorescent protein (GFP), enabling researchers to track their location.
- Combining spectral variants of GFP and RFP provides simultaneous visualization of multiple proteins.
- Confocal microscopy uses a laser to illuminate and detect light from the sample, creating sharper images by reducing out-of-focus light. Image data are assembled from optical sections.
Contrast Techniques
- Various techniques enhance contrast in microscopy, such as staining, phase contrast, and differential interference contrast (DIC).
- Different staining methods generate contrast by coloring specific structures, improving visibility.
- Phase contrast and DIC are optical techniques that use different refractive indices to distinguish varying density areas inside cells.
- Negative staining darkens the background around samples, making particles or molecules stand out.
Preparing Specimens for Microscopy
- Preparing specimens involves a combination of steps such as fixation, dehydration, and embedding to preserve cells and tissues.
- Fixatives like formaldehyde and glutaraldehyde are used to preserve cellular structure by immobilizing proteins and structures.
- Dehydration removes water from specimens, allowing them to be infiltrated with embedding media.
- Embedding media, such as wax or plastic, surround the specimen to create a rigid block, aiding in sectioning.
- Sectioning with a microtome generates thin slices of specimens.
- Sections are mounted on microscope slides followed by staining to generate contrast.
Image Resolution
- Magnification increases the size of an image, but resolution is the ability to distinguish two separate points in an image.
- Magnification without sufficient resolution can lead to empty magnification, without improving clarity.
- The resolution of a light microscope is limited by the wavelength of light, typically to about 0.2 µm.
- Electron microscopes, using electrons instead of light, offer significantly higher resolution, allowing visualization of structures in the nanometer range (0.2 nm).
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Description
Explore various microscopy techniques that enable visualization of structures at different scales, from atoms to cells. This quiz covers light microscopy, electron microscopy, and advanced methods like atomic force microscopy and fluorescence microscopy. Test your knowledge on how these techniques enhance our understanding of biological materials.
