5. Light Microscopy (LM) Types

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TAMU BMEN 311 - Imaging Living Systems [Light Microscopy Types]

16 Terms

1

Contrast enhancement

  • distinguish fine structural details and visualize transparent or low-contrast specimens more clearly.

  • improves visibility of transparent features

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2

specimans examined in LM

  • biological tissues and cells

  • transparent microorganisms (bacteria and protozoa)

  • cellular components like organelles

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3

Types of Light Microscopy

  • Brightfield*

  • Darkfield

  • Phase Contrast

  • Differential Interference Contrast (DIC)

  • Polarized Light

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4

BRIGHTFIELD

  • standard lab microscope

  • produces image on a bright background

  • Cells and tissues have intrinsically low contrast when imaged in brightfield mode

  • using histology stains to improve contrast = toxic → KILLS CELLS

<ul><li><p>standard lab microscope</p></li><li><p>produces image on a bright background</p></li><li><p>Cells and tissues have intrinsically low contrast when imaged in brightfield mode </p></li><li><p>using histology stains to improve contrast = toxic → KILLS CELLS</p></li></ul><p></p>
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5

Brightfield applications

routine examination of blood smears, histology, and cytology studies

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6

DARKFIELD

  • stars become visible at night

  • Increases contrast without staining

  • producing a bright image on a darker background

  • useful for viewing THIN, LIVE specimens

  • sensitive to debris

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7

Darkfield applications

  • image syphilis

  • studying external details

  • tiny diffracting objects like bacteria and isolated organelles

<ul><li><p><strong>image syphilis</strong></p></li><li><p><u>studying external details</u> </p></li><li><p>tiny diffracting objects like bacteria and isolated organelles</p></li></ul><p></p>
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8

PHASE CONTRAST

  • uses refraction and interference caused by speciman’s structures create high-contrast, high-resolution images

  • no staining, useful for viewing live specimens.

  • utilizes phase shifts caused by differences in refractive indices to enhance contrast in transparent, thin specimens.

  • subject to “halo” artifacts

<ul><li><p>uses refraction and interference caused by speciman’s structures create high-contrast, high-resolution images </p></li><li><p>no staining, useful for viewing live specimens. </p></li><li><p><strong><u>utilizes phase shifts caused by differences in refractive indices </u></strong>to enhance contrast in <strong><em>transparent, thin </em></strong>specimens. </p></li><li><p>subject to “halo” artifacts</p></li></ul><p></p>
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9

phase contrast applications

nuclei, neurons membranes, cell division, tissue engineering applications

<p>nuclei, neurons membranes, cell division, tissue engineering applications</p>
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10
<p>Label the image </p>

Label the image

  1. brightfield

  2. dark field

  3. phase contrast

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11

DIFFERENTIAL INTERFERENCE CONTRAST (DIC)

  • interference patterns → enhance contrast = high-contrast images of living organisms with (pseudo) 3D appearance

  • useful in distinguishing structures

  • enhances contrast in specimens with slight variations in thickness and refractive index

    • USEFUL FOR THINK SPECIMANS

    • NO “halo” artifacts

    • expensive

<ul><li><p>interference patterns → enhance contrast = high-contrast images of <u>living </u>organisms with (pseudo) 3D appearance</p></li><li><p>useful in distinguishing structures</p></li><li><p>enhances contrast in specimens with slight variations in thickness and refractive index</p><ul><li><p>USEFUL FOR THINK SPECIMANS</p></li><li><p>NO “halo” artifacts</p></li><li><p>expensive</p></li></ul></li></ul><p></p>
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12

DIC Applications

cell motility, cellular structure, cell division (pic), tissue engineering applications

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13

POLARIZING LIGHT

polarized light and filter to study birefringent specimens

<p>polarized light and filter to study birefringent specimens</p>
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14

Birefringence

optical property of a material

  • refractive index depends on polarization and propagation direction of light. (aka double refraction)

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15

Polarizing Light Applications

spindle and collagen fibers (pic), gout crystals, (normal and malaria-infected) red blood cells; optically anisotropic specimens (birefringent)

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16

light polarization

knowt flashcard image
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