User:Wilfred J. Poppinga/Notebook/cAMP compartmentalization/2010/08/13

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 * style="background-color: #EEE"|[[Image:owwnotebook_icon.png|128px]] SDS-PAGE Lysates, strip blot
 * style="background-color: #F2F2F2" align="center"|  |Main project page
 * style="background-color: #F2F2F2" align="center"|  |Main project page


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SDS-PAGE

 * Lysate samples were prepared, undiluted samples (75 µL) with NuPage loading buffer + DTT (25 µL)
 * Samples were put to 70 °C for 10 min.
 * Samples were put to 4-12% Gradient NuPage gel 200 V

Stripping blot

 * Stripping solution
 * 50 mL dH2O
 * 5 mL Tris-Cl (1.0 M, pH 6.8) (Stacking gel buffer SDS-PAGE)
 * 1 g SDS
 * 150 mg DTT (Add later)
 * 1) Prewarm buffer (w/o DTT) for 30 min. to 70 °C in waterbath
 * 2) Add DTT just before adding membrane
 * 3) Completely immerse membrane
 * 4) Incubate 30 min @ 70 °C
 * 5) Wash 3x with dH2O or TBS-T
 * 6) Membrane can now be blocked again

Results

 * Gel after blotting


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