User:Karmella Haynes/Notebook/PcTF Genomics/2013/06/18: Difference between revisions
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<u>Luc assay</u> | <u>Luc assay</u> | ||
* Filtered 500 μL cells through strainer caps (<font color="red">should use | * Filtered 500 μL cells through strainer caps (<font color="red">should use 600 μL next time</font>) | ||
* Transferred 100 μL strained cells to 3 wells in 96-well plate (black walls; clear, flat bottom) | * Transferred 100 μL strained cells to 3 wells in 96-well plate (black walls; clear, flat bottom) | ||
** Saved remainder of strained cells for flow cytometry/ counting | ** Saved remainder of strained cells for flow cytometry/ counting | ||
Line 38: | Line 38: | ||
* QUICKLY Added 100 μL of complete steady-luc buffer to each sample (pipetted up/down once to mix) | * QUICKLY Added 100 μL of complete steady-luc buffer to each sample (pipetted up/down once to mix) | ||
* Incubated at room temp/ 10 min. | * Incubated at room temp/ 10 min. | ||
* Synergy H1 reader - Created new protocol: KAH_luciferase | |||
** Optimized specs: Luminescence, Endpoint, Full plate, Integration time 0:01.00 (MM:SS.ss), Filter set 1, Emission: full light, Optics: top, '''Gain 200''', Read Speed: normal, Delay: 100 msec, Extended Dynamic Range, Read Height: 1 mm | |||
<u>Cell | <u>Cell counts</u> | ||
* Wang lab's Accuri flow cytometer. Set machine to | * Wang lab's Accuri flow cytometer. | ||
* Set machine to read 20 uL of cells | |||
* Be sure to "clean" with water-run in between samples | |||
{| class="wikitable" border="0" cellspacing="3" <!-- Rxn. table --> | {| class="wikitable" border="0" cellspacing="3" <!-- Rxn. table --> | ||
|-valign="top" | |-valign="top" | ||
| <u> | | <u>Sample ID</u> || <u>Gated count/ 20 μL</u> || || <u>Cells/ 100 μL</u> | ||
|- | |- | ||
| | | sample 1 || 8077 || x5 = || 40,385 | ||
|- | |- | ||
| | | sample 2 || 9552 || x5 = || 47,760 | ||
|- | |- | ||
| | | sample 3 || 9460 || x5 = || 47,300 | ||
|- | |- | ||
| | | sample 4 || 3608 || x5 = || 18,040 | ||
|- | |- | ||
| | | sample 5 || 4723 || x5 = || 23,615 | ||
|- | |- | ||
| | | sample 6 || 5851 || x5 = || 29,255 | ||
|} | |} | ||
Revision as of 19:44, 18 June 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> </html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html> | ||||||||||||||||||||||||||||
06/18/13
Luciferase activity assay - time point 1
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