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  1. Methods of fixing
    • heat-fix
    • chemically fix (95% methanol)
  2. Simple stain
    • Usually one stain
    • Positive ion (basic dye) stains cell (direct)
    • Negative ion (acidic dye) stains background (negative) 
  3. Gram staining order/purpose of each step
    • 1. crystal violet (primary stain)
    • 2. iodine (mordant)
    • 3. ehtanol (decolorizing agent)
    • 4. safranin (counterstain)
    • Gram-positive (purple): much peptioglycan in cell wallsGram-negative (pink): thin layer of peptidoglycan surrounded by lipoproteins, phospholipids, and lipopolysaccharides.
  4. Acid-Fast Staining
    • Differential stain
    • Cell walls of acid-fast organisms contain mycolic acid
    • Smear is flooded with carbolfuchsin
    • Smear is heated
    • Washed with acid-alcohol mixture (decolorizer)
    • Methylene blue (counterstain)
  5. Endospore stain
    • Endospores are impermeable to most stains
    • Heat applied to drive the stain in.
    • Malachite green onto absorbant paper over smear
    • Steam for 5 minutes
    • Wash with water
    • Cover with safranin for 30 seconds
    • Wash with water
  6. Capsule stain
    • Simple stains won't adhere due to capsule's non-ionic nature
    • Most stain the background and cell, leaving the capsule unstained
    • Congo red on slide
    • Bacteria into congo red
    • Allow to dry
    • Cover with acid-alcohol
    • Wash smears with water
    • Cover with acid fuchsin
    • Wash smears with water
  7. Flagella stain
    • Peritrichous (all over)
    • Monotrichous (single at one end)
    • Lophotrichous (much at one end)
    • Amphitrichous (at both ends)
    • Distilled water + bacteria on slide
    • Allow drop to flow to other side (tilt)
    • Let air dry
    • Cover with flagella stain for 4 minutes
    • Rinse stain with water
Card Set:
2012-09-21 20:44:22

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