User:Wilfred J. Poppinga/Notebook/cAMP compartmentalization/2010/07/10: Difference between revisions

From OpenWetWare
Jump to navigationJump to search
Line 345: Line 345:
* Ponceau staining of AKAP250 Co-IP blot
* Ponceau staining of AKAP250 Co-IP blot
[[Image:10072010_Ponceau.png|500px]]
[[Image:10072010_Ponceau.png|500px]]
[[Image:10072010_coomassie.png|500px]]
==Discussion==
==Discussion==
* Very peculiar that the Ponceau staining gives the exact same image as the cAMP precipitations.  
* Very peculiar that the Ponceau staining gives the exact same image as the cAMP precipitations.  

Revision as of 08:57, 10 July 2010

<html><style type="text/css"> .todo { color: red } .done { color: green} </style></html>

Project name <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>

Summary

  • Destaining gel remains that was coomassie stained after blotting the cAMP precipitation samples
  • Putting the AKAP250 immunoprecipitation samples to gel and blot them
  • Gradient (4-12%) SDS-PAGE (NuPage) for cAMP Precipitations
Gel loading hTERT D9 + HEK293 & HeLa -S see codes 21June2010
# 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15
Gel
cAMP prec. 		X X Invitrogen Marker
(8 μL) 		HeLa -S
(20 μL) 		HEK293
(20 μL) 		Biorad Marker
(8 μL) 		C1
(20 μL) 		C2
(20 μL) 		C3
(20 μL) 		S1
(20 μL) 		S2
(20 μL) 		S3
(20 μL) 		X X X
  • After this gel has been blot it will be blocked and then together with the other blot treated with antiAKAP250 antibody

Notes

  • During wash steps of co-ip centrifuge steps were performed at 4400 rpm instead of 2500 rpm until the third wash step
  • Supernatant of ON incubation for co-ip was saved, "NAIP 250 9.7.10" stored @ -20 °C
  • Made new MES NuPage buffer

Results

  • Coomassie staining was put together with the ponceau S staining of yesterday
  • Ponceau staining of AKAP250 Co-IP blot

Discussion

  • Very peculiar that the Ponceau staining gives the exact same image as the cAMP precipitations.


Retrieved from "https://openwetware.org/mediawiki/index.php?title=User:Wilfred_J._Poppinga/Notebook/cAMP_compartmentalization/2010/07/10&oldid=431492"