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Published byTerence Carpenter Modified over 9 years ago
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MICROSCOPY AND STAINING
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Leeuwenhoek’s Investigation 2
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3 Metric Units
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4 Light Properties Wavelength http://eyetricks.com/0101.htm
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5 Light Properties Reflection Transmission
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6 Light Microscopy Types Compound Bright Field
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7 Light Properties Resolution
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8 Wavelength/Resolution Interaction
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9 Light Properties Absorption Refraction http://interactagram.com/physics/optics/refraction/
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Effect of Immersion Oil 10
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Disappearing Diamond 11
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Diffraction 12
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Diffraction 13
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14 Microscopy — Dark Field
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15 Microscopy — Phase Contrast Dual Beam
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16 Microscopy — DIC (Nomarsky) Differential Interference Contrast
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17 Microscopy — Fluorescence
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18 Microscopy — Confocal Confocal
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19 Microscopy Imaging Digital
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Same Image – Different Techniques 20
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21 Electron Microscopy Transmission (TEM or EM) Scanning (SEM) Scanning Tunneling (STM) Atomic Force (AFM) Scanning Acoustic (SAM) http://micro.magnet.fsu.edu/primer/ virtual/virtual.html *animation
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Freeze - fracture 22
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SEM Shadow Casting 23
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SEM freeze-etch 24
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25 Electron Microscopy Images
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STM 27
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AFM 28
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29 Microscopy Techniques Wet Mounts Smears Drop
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Simple Stain 30 Provide basic contrast for light compound microscope Anionic = acidic dyes: eosin picric acid Cationic = basic dyes: methylene blue crystal violet safranin malachite green
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Differential Stain 31
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Differential Stain 32
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33 1. Gram Stain Crystal violet stain Iodine mordant 95% ethanol decolorizer or ethanol/acetone Safranin counterstain
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34 2. Acid Fast Stain (Ziehl-Neelson) Paul Ehrlich in 1882; Detecting tuberculosis and leprosy – causing organisms. Carbolfuchsin stain 3% HCl declorizer in 95% ethanol Methylene Blue counterstain
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35 3. Special Stain: Negative Stain Cell size Cell shape Indian Ink or acidic dyes for background Crystal Violet or Methylene Blue for counterstain
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36 4. Special Stain: Flagellar Stain Silver; Very difficult and time consuming;
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37 5. Special Stain: Endospore Stain (Schaeffer-Fulton) Heat-fix Malachite Green stain Steam for 5 min to make the endo- spore wall permeable Rinse in water for 30 sec Decolor Safranin counterstain
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