Griffitts:Bacterial Staining

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Solutions

  • LB Broth
  • 0.85% NaCl
  • 20 mM propidium iodide (in kit in freezer)
  • 3.34 mM SYTO 9 (in kit in freezer)

Procedure

  • Grow the bacterial cells to an OD600 of 0.5
    • If the OD600 is > 0.5, dilute appropriately
  • Pellet at 13,200 rpm for 1 minute
  • Resuspend in 1 mL 0.85% NaCl
  • Incubate 10 minutes
  • Pellet at 13,200 rpm for 1 minute
  • Resuspend in 500 μL 0.85% NaCl
  • Add 0.75 μL of 20 mM propidium iodide
  • Add 0.75 μL of 3.34 mM SYTO 9
  • Incubate in the dark at room temperature for 15 minutes
  • Proceed to fluorescent microscopy
    • PI (red) stains dead cells
    • SYTO 9 (green) stains live cells
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