Imaging Techniques in Microscopy

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The ______ microscopy uses fluorescent dyes to produce an image of the sample.

fluorescence

The ______ lenses collect and focus light from the sample.

objective

The process of ______ preserves the sample by cross-linking proteins.

fixation

The ______ staining uses a dye to highlight specific structures or features.

positive

The ______ microscopy uses a beam of electrons to produce an image of the sample.

electron

The ______ mechanism allows the user to adjust the focus of the image.

focusing

The ______ microscopy uses a physical probe to produce an image of the sample's surface.

scanning probe

The ______ microscopy uses a laser to produce a high-resolution image of the sample.

confocal

Study Notes

Imaging Techniques

  • Brightfield microscopy: uses visible light to produce a magnified image of the sample
  • Fluorescence microscopy: uses fluorescent dyes to produce an image of the sample
  • Phase contrast microscopy: uses differences in refractive indices to produce an image of the sample
  • Darkfield microscopy: uses scattered light to produce an image of the sample
  • Confocal microscopy: uses a laser to produce a high-resolution image of the sample
  • Super-resolution microscopy: uses techniques such as STORM or STED to produce an image of the sample with resolution below the diffraction limit

Microscope Components

  • Objective lenses: collect and focus light from the sample
  • Ocular lenses: magnify the image formed by the objective lenses
  • Stage: holds the sample in place
  • Illumination system: provides light to the sample
  • Focusing mechanism: allows the user to adjust the focus of the image

Sample Preparation

  • Fixation: preserves the sample by cross-linking proteins
  • Dehydration: removes water from the sample to prevent distortion
  • Embedding: surrounds the sample with a medium to support it
  • Sectioning: cuts the sample into thin slices
  • Mounting: attaches the sample to a slide or other support

Staining

  • Positive staining: uses a dye to highlight specific structures or features
  • Negative staining: uses a dye to highlight the background or surrounding material
  • Special staining: uses specific dyes to highlight specific structures or features
  • Immunofluorescence: uses antibodies to label specific structures or features

Microscopy Types

  • Light microscopy: uses visible light to produce an image of the sample
  • Electron microscopy: uses a beam of electrons to produce an image of the sample
    • Transmission electron microscopy (TEM): uses a beam of electrons to produce an image of the sample
    • Scanning electron microscopy (SEM): uses a beam of electrons to produce an image of the sample's surface
  • Scanning probe microscopy: uses a physical probe to produce an image of the sample's surface
    • Atomic force microscopy (AFM): uses a physical probe to produce an image of the sample's surface
    • Scanning tunneling microscopy (STM): uses a physical probe to produce an image of the sample's surface

Imaging Techniques

  • Brightfield microscopy uses visible light to produce a magnified image of the sample, with the sample appearing darker than the background.
  • Fluorescence microscopy uses fluorescent dyes to produce an image of the sample, with the dyed structures emitting light at specific wavelengths.
  • Phase contrast microscopy uses differences in refractive indices to produce an image of the sample, with the contrast created by the variation in refractive indices.
  • Darkfield microscopy uses scattered light to produce an image of the sample, with the sample appearing bright against a dark background.
  • Confocal microscopy uses a laser to produce a high-resolution image of the sample, with the laser beam focused onto a specific point on the sample.
  • Super-resolution microscopy uses techniques such as STORM or STED to produce an image of the sample with resolution below the diffraction limit, allowing for detailed imaging of structures smaller than the wavelength of light.

Microscope Components

  • Objective lenses collect and focus light from the sample, with the objective lens being the primary lens that collects and focuses the light.
  • Ocular lenses magnify the image formed by the objective lenses, with the ocular lens being responsible for further magnifying the image.
  • Stage holds the sample in place, allowing for precise positioning and control of the sample.
  • Illumination system provides light to the sample, with the type of illumination system used depending on the type of microscopy being performed.
  • Focusing mechanism allows the user to adjust the focus of the image, with the focusing mechanism typically consisting of a coarse adjustment knob and a fine adjustment knob.

Sample Preparation

  • Fixation preserves the sample by cross-linking proteins, preventing degradation and maintaining the sample's structure.
  • Dehydration removes water from the sample to prevent distortion, with the sample being dehydrated using a series of solvents of increasing concentration.
  • Embedding surrounds the sample with a medium to support it, with the medium used depending on the type of microscopy being performed.
  • Sectioning cuts the sample into thin slices, allowing for detailed imaging of the sample's internal structures.
  • Mounting attaches the sample to a slide or other support, allowing for easy handling and manipulation of the sample.

Staining

  • Positive staining uses a dye to highlight specific structures or features, with the dye binding to specific molecules or structures.
  • Negative staining uses a dye to highlight the background or surrounding material, with the dye binding to everything except the structure of interest.
  • Special staining uses specific dyes to highlight specific structures or features, with the choice of dye depending on the type of sample and the structure being studied.
  • Immunofluorescence uses antibodies to label specific structures or features, with the antibodies binding to specific molecules or structures and emitting light at specific wavelengths.

Microscopy Types

  • Light microscopy uses visible light to produce an image of the sample, with the sample being illuminated by a light source.
  • Electron microscopy uses a beam of electrons to produce an image of the sample, with the electron beam being focused onto the sample.
    • Transmission electron microscopy (TEM) uses a beam of electrons to produce an image of the sample, with the electrons passing through the sample.
    • Scanning electron microscopy (SEM) uses a beam of electrons to produce an image of the sample's surface, with the electrons being scanned across the sample's surface.
  • Scanning probe microscopy uses a physical probe to produce an image of the sample's surface, with the probe being scanned across the sample's surface.
    • Atomic force microscopy (AFM) uses a physical probe to produce an image of the sample's surface, with the probe measuring the deflection of the cantilever.
    • Scanning tunneling microscopy (STM) uses a physical probe to produce an image of the sample's surface, with the probe measuring the tunneling current between the probe and the sample.

This quiz covers different types of microscopy techniques used to produce magnified images of samples. Learn about brightfield, fluorescence, phase contrast, darkfield, confocal, and super-resolution microscopy.

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