User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/18: Difference between revisions
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'''Luciferase activity assay - time point 1'''<br> | '''Luciferase activity assay - time point 1'''<br> | ||
<u>Cell prep</u> | |||
* Harvested cells | * Harvested cells | ||
** Seeded new dox+ plate with 1/2 of cells | ** Seeded new dox+ plate with 1/2 of cells | ||
** Pelleted other half, resuspended in 2 mL FACS buffer | ** Pelleted other half, resuspended in 2 mL FACS buffer | ||
<u>Assay reagents</u> | |||
* Used the Biotium Steady-Luc Firefly HTS Assay Kit | * 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)''' | * 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''' | * 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 dH<sub>2</sub>O to the vial | ||
* Made 2000 μL complete steady-luc buffer: Add 40 μL 50x D-luciferin to 2 mL buffer | |||
<u>Luc assay</u> | |||
* | <u>Cell count</u> | ||
* Wang lab's Accuri flow cytometer. Set machine to sample 2 uL of cells | |||
Revision as of 17:54, 18 June 2013
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06/18/13
Luciferase activity assay - time point 1
Luc assay Cell count
--> Reaction conditions
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