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

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Gravin Co-IP gel Main project page
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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 AKAP250 Co-IP 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
Gravin Co-IP
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.


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