Difference between revisions of "User:Karmella Haynes/Notebook/PcTF Genomics/2013/07/04"

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(mm/dd/yy)
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==mm/dd/yy==
 
==mm/dd/yy==
 
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* Gl4-EED/luc time/dose - luc assay & cell counts
  
 
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<u>Luc assay</u>
 
<u>Luc assay</u>
 
* Filtered '''700 μL cells''' through strainer caps  
 
* Filtered '''700 μL cells''' through strainer caps  
 +
* Used opaque white Costar plate (from Rege lab)
 +
* Samples loaded in triplicate (by columns)
 +
* Included luc buffer + FACS-buffer "blank" sample (well D1)
 
* Other steps same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/07/04 6/18/13]
 
* Other steps same as [http://openwetware.org/wiki/User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/07/04 6/18/13]
  
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| <u>Sample ID</u> || <u>Gated count/ 20 μL</u> || &nbsp; || <u>Cells/ 100 μL</u>
 
| <u>Sample ID</u> || <u>Gated count/ 20 μL</u> || &nbsp; || <u>Cells/ 100 μL</u>
 
|-
 
|-
| sample 1 || 10,880 || x5 = || 54,400
+
| sample 1 || ### || x5 = || 54,400
 
|-
 
|-
 
| sample 2 || 10,430 || x5 = || 52,150
 
| sample 2 || 10,430 || x5 = || 52,150

Revision as of 20:14, 4 July 2013

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mm/dd/yy

  • Gl4-EED/luc time/dose - luc assay & cell counts

Luciferase activity assay - time point: 4 days


Cell prep

  • Harvested cells (induced on 6/30/13)
    • Seeded new 2 new plates with non-induced cells from 6/30/13
    • Pelleted all of the dox+ cells, resuspended in 2 mL FACS buffer


Assay reagents

  • Used the Biotium Steady-Luc Firefly HTS Assay Kit (same as 6/18/13)


Luc assay

  • Filtered 700 μL cells through strainer caps
  • Used opaque white Costar plate (from Rege lab)
  • Samples loaded in triplicate (by columns)
  • Included luc buffer + FACS-buffer "blank" sample (well D1)
  • Other steps same as 6/18/13


Cell counts

  • Wang lab's Accuri flow cytometer.
  • Set machine to read 20 uL of cells
  • Be sure to "clean" with water-run in between samples


Sample ID Gated count/ 20 μL   Cells/ 100 μL
sample 1 ### x5 = 54,400
sample 2 10,430 x5 = 52,150
sample 3 11,897 x5 = 59,485
sample 4 10,316 x5 = 51,580
sample 5 9,784 x5 = 48,920
sample 6 11,549 x5 = 57,745