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1 1 Lec 5 Chapter 4 Microscopy and Stains: Cell Size and Morphology Sept 3, 2008 Office phone: 231- 9708 LS-1 room 215 2 Anton van Leeuwenhoek: “There are more animals living in the scum on the teeth in a man’s mouth than there are in a whole kingdom.” Resolution 200nm 3 Refractive Index Measure of light-bending ability of the medium. Changing the refractive index of the specimen relative to the medium increases the contrast between the two. Staining is used to change the refractive index.
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2 4 Immersion Oil • Air has a different refractive index than glass. • Most light would be lost if it had to travel through air between the specimen and the objective lens. • Immersion oil has about the same refractive index as glass and increases the resolving power of the lens. 5 Resolution and Wavelength • The ability of the lens to distinguish two points a specified distance apart. ¾ Function of both the wavelength of light used to image and the light gathering ability of the objective lens being used (why oil helps) • The shorter the wavelength, the greater the resolution. • Best resolution of light microscopes is 0.2 μ m (2,000X) • Electron microscopes resolution 0.2 to 0.5nm 6 Staining for Light Microscopy Utilization of stains on specimens can introduce artifacts and distortion due to the chemical influences on cell structure (dehydration for example) Simple stains – increase contrast –use one dye to highlight cell morphology ¾ Basic dyes + charged ### ¾ Acid dyes - charged Differential stains – differentiate between organisms or structures by using a primary dye and a contrasting counterstain to distinguish cell types or parts ¾ Gram stain – differentiates between two kinds of cell wall structures
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This note was uploaded on 02/11/2009 for the course BIOL 2604 taught by Professor Sldaniel during the Spring '08 term at Virginia Tech.

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