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Project name
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Summary
Materials & Methods
Materials
Method
- Blocking blot of 21August2010 for pre-IgG antibodies
- Blocking occured ~8 hours @ RT
- Rabbit Pre-Immune serum taken just before immunisation was used(ZDE10030 animal 122 antibody (Eurogentech antibody service)) 1:1000 and incubated ON @ 4°C
- SDS-PAGE of cAMP precipitation samples 21June2010
- 10 min. 70 °C → RT
- 5 min. 95 °C (no DTT in this loading bufer) → RT
- 1 min. 95 °C
- 200 V
4-12% Gradient NuPage Gel loading hTERT D9 + HEK293 & HeLa -S see codes 21June2010
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1
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8
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9
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10
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Gel cAMP prec.
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Invitrogen Marker (8 μL)
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HeLa -S (20 μL)
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HEK293 (10 μL)
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Biorad Marker (8 μL)
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C1 (20 μL)
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C2 (20 μL)
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C3 (20 μL)
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S1 (<20 μL)
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S2 (<20 μL)
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S3 (<20 μL)
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- Western blot, 2h @ 30 V
- Put HEK293 (p17) to S0
- Weighed AKAP18δ-L314E (3.21 mg) and dissolved it (111 μL DMSO) for used in RII overlay
Notes
- SDS-PAGE
- Gel was burned?? after 1h 13 min. and due to current change machine stopped running
- SDS-PAGE was stopped at 2 h 2 min., burning discoloring was spreading
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