1. Aspirate off media.
  2. Rinse wells with 500uL of PBS/versene. Aspirate.
  3. Add 200uL of trypsin/EDTA to each well. Incubate at room temperature for 90 seconds, then neutralize with 800uL of warm complete media
  4. Triturate cells to remove clumps (pipette up and down)
  5. Transfer suspended cells to 15 mL Falcon Tubes.
  6. Centrifuge cells 5min 2500 rpm at 23°C.
  7. Resuspend cells in warm PBS, 10% FBS, 1% sodium azide to a concentration of 5X10^6 cells per mL with a total volume of 1000 uL
    NOTE: DO NOT ASPIRATE SOLUTIONS CONTAINING SODIUM AZIDE. Pipette waste into an empty bottle and dispose of it appropriately later. 
  8. Add Beta Amyloid at 500 nM concentration (or other)
  9. Incubate at 37C for 1hr 30min
  10. Move all warm reagents to the refrigerator
  11. Move to cold room for 30 min
  12. Centrifuge cells 5min 2500 rpm at 4°C
  13. Resuspend cells in 1000 uL ice-cold PBS, 10% FBS, 1% sodium azide
  14. Add Streptavidin to a 10 ug/mL (5 uL for our aliquot)
  15. Incubate in the cold room in the dark for 1hr 30 min
  16. Centrifuge cells 5min 2500 rpm 4°C
  17. Resuspend cells in 1000 uL of ice cold PBS, 10% FBS, 1% sodium azide
  18. Count cells on a hemacytometer (not all of samples, just a few.)  Dilute to ~1e6 cells/ml with ice-cold  PBS, 10% FBS, 1% sodium azide. 
  19. Transfer suspension to cytometry tubes.
  20. Run flow cytometry.
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