User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/18

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06/18/13

  • Luciferase activity assay



Luciferase activity assay - time point 1


Cell prep

  • Harvested cells
    • Seeded new dox+ plate with 1/2 of cells
    • Pelleted other half, resuspended in 2 mL FACS buffer


Assay reagents

  • Used the Biotium Steady-Luc Firefly HTS Assay Kit
  • Thawed reagents to room temp
  • 6 experimental samples x 3 repliates = 18 assays (one time point)
  • Need 100 μL complete steady-luc buffer per assay = 1800 μL complete steady-luc buffer
  • Dissolved dry D-luciferin to make 50x stock by adding 80 μL dH2O to the vial
  • Made 2000 μL complete steady-luc buffer: Add 40 μL 50x D-luciferin to 2 mL buffer

Luc assay

Cell count

  • Wang lab's Accuri flow cytometer. Set machine to sample 2 uL of cells


Reagent Volume
reagent 1 # μL
reagent 2 #
reagent 3 #
reagent 4 #
dH2O #
  # μL

--> Reaction conditions



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