User:Karmella Haynes/Notebook/PcTF Genomics/2013/07/06: Difference between revisions

From OpenWetWare
Jump to navigationJump to search
Line 10: Line 10:
==07/06/13==
==07/06/13==
<!-- Precede finished items with a checkmark &#x2713; -->
<!-- Precede finished items with a checkmark &#x2713; -->
* Line item 1
* Gal4-EED/luc dox time point (48 hours)
* Gal4-EED/luc PcTF transfection - microscopy & flow cytometry




----
----
'''Luciferase activity assay - time point: 4 days'''<br>
'''Luciferase activity assay - time point: 2 days'''<br>
 


<u>Cell prep</u>
<u>Cell prep</u>

Revision as of 15:05, 6 July 2013

Pc-TF Genomics <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>


07/06/13

  • Gal4-EED/luc dox time point (48 hours)
  • Gal4-EED/luc PcTF transfection - microscopy & flow cytometry


Luciferase activity assay - time point: 2 days

Cell prep

  • Harvested cells (induced on 7/04/13)
    • Seeded new 2 new plates with non-induced cells from 7/04/13
    • 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 13,844 x5 = 69,220
sample 2 12,286 x5 = 61,430
sample 3 15,784 x5 = 78,920
sample 4 16,323 x5 = 81,615
sample 5 16,545 x5 = 82,725
sample 6 17,885 x5 = 89,425



Retrieved from "https://openwetware.org/mediawiki/index.php?title=User:Karmella_Haynes/Notebook/PcTF_Genomics/2013/07/06&oldid=707514"