Staining Sections for Light Microscopy

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Staining Sections for Light Microscopy
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Why stain?

• What and where is it?
• How much/how many are there?

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Why does anything stain?

• Affinity
• electrostatic (ionic, hydrogen bonding)
• covalent
• Van der Waals
• Non-affinity
• vital staining
• negative staining
• catalytic staining

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Why doesnt everything stain?

• Selectivity
• Sensitivity

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Some common stains

• Toluidine blue
• metachromatic--color depends on how much of the
  dye is aggregated
• H E
• acid dye (eosin) and basic dye (hematoxylin)

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Mallorys Trichrome Stain for collagen
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General differential staining vs histochemistry
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Flow cytometry
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• Histochemistry vs cytochemistry

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The PAS Reaction(Periodic-acid Schiff)

• Non-specifically stains carbohydrates
• Used to demonstrate glycogen,mucin, etc.
• Binding is stoichiometric
• Can also be used to quantify DNA, other things

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How it works

• Periodic acid oxidizes free end of reducing
  sugars to aldehydes
• Schiff reagent covalently binds to the aldehydes

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Controls?
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Stem stained with toluidine blue
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Quantitation using flow cytometry