MICROSCOPES Microscopic Instruments differ in their lenses and the source of their illumination.
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Transcript of MICROSCOPES Microscopic Instruments differ in their lenses and the source of their illumination.
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MICROSCOPES
Microscopic Instruments differ in their lenses and the source of their illumination
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Microscopes
• All microscopes operate on the same basic principles– Energy is projected toward an object (MO)– Energy bounces off of the object and
creates an impression on a sensing device– The device can be a TV screen, a piece of
film or the human eye– The image reveals the form, shape, size
and structural features of the object
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I. LIGHT MICROSCOPE
Visible light illuminates the object
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A. STEREO MICROSCOPE (SM)
• Light bulb for a light source
• Magnification of 35X
• Surface picture (3D)
Spore under a Stereo Microscope
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B. COMPOUND MICROSCOPE (CM)
• Most commonly used – 2 lenses
• Ocular lens – 10X
• Objective lenses
• 10X, 40X, 100X
• Some can even go to
2000X
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B. COMPOUND MICROSCOPE (CM)
• Light bulb for light source
• Light goes through object
• Small or thin slice object
• Size and shape of bacteria cell can be seen
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II. ELECTRON MICROSCOPE
Thousand times better than a light
microscope.
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II. ELECTRON MICROSCOPE
• German physicist Ernst Ruska
• Showed that electrons can flow in a sealed tube if a vacuum is maintained
• Magnets can be used to
pinpoint the flow of
electrons onto
an object
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II. ELECTRON MICROSCOPE
• Depending upon the density of structures in the object, the electrons are either absorbed or deflected
• The electrons form an image that can be projected onto a screen and outlines the structures in the object
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A. TRANSMISSION ELECTRON MICROSCOPE (TEM) 1931
• Used to see internal cell structures in detail
• Ultrathin sections of a specimen must be prepared because the electron beam can only penetrate matter a short distance
• Sections are floated in water and picked up on a wire grid
• Sections are inserted into the vacuum chamber of the microscope
• Electrons illuminate the specimen
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A. TRANSMISSION ELECTRON MICROSCOPE (TEM) 1931
• Magnetic field focuses the beam of electrons (as condenser lens of light
• No ocular lens, electrons hit electron sensitive screen to create image
• 200,000X – Strongest
Microscope Transmission electron micrograph of E. coli
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B. SCANNING ELECTRON MICROSCOPE (SEM) 1960’s
• Permits the surface of objects to be seen without having to make thin sections
• Specimens are placed in the vacuum chamber and coated with a thin layer of gold to increase conductivity
• The electron beam sweeps across the object and knocks loose showers of electrons
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B. SCANNING ELECTRON MICROSCOPE (SEM) 1960’s
• A 3D image builds line by line like a TV receiver – can see surface detail
• Can magnify 50X – 100,000X
• See image on TV screen
SEM 3D Photo of E. Coli
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B. SCANNING ELECTRON MICROSCOPE (SEM) 1960’s
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III. VARIATIONS
Manipulation of Light
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A. DARK-FIELD MICROSCOPE
• Highlights specimen against a black background – only object is illuminated
• Light scattered and hits object from different angles
• Like us seeing the moon at night because sunlight from behind the earth reflects off the moon
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A. DARK-FIELD MICROSCOPE
• Good for living MOs to see size, shape and motility
• Helps in diagnosis of some diseases caused by spiral bacteria because they are so small
• Treponema palladum causes syhphilis and is identified from scrapings of the infected lesions
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B. PHASE CONTRAST MICROSCOPE• Used in research labs for observing living MO
and their movement in medium where they are growing – no staining
• Same magnification as compound microscope but it detects small differences in density
• Compound microscope that increases the contrast between denser MOs and surrounding medium (MO denser than medium)
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B. PHASE CONTRAST MICROSCOPE
Epithelial Cells - Phase Contrast Microscope Epithelial Cells -
Compound Microscope
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C. FLUORESCENCE MICROSCOPE
• MO’s are coated with fluorescent dye and illuminated with UV light
• Coated MO’s appear to fluoresce
• Applications in
medical microbiology
tagged antibodies
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D. SCANNING TUNNELING MICROSCOPE (STM) 1981
• Focus on surface of object, produce a map showing bumps and valleys of atoms
• Map surface as blind person with a cane (tap)
• No special prep to sample
Surface of Blue NickelSurface of CopperSurface of Blue Platinum
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