|0.25 M Tris
1.92 M Glycin
|Working solution:||Dilute 100 ml of 10x concentrate with 200 ml methanol and
700 ml distilled water.
- Cut transfer membrane and four pieces Whatmann 3 MM paper to gel size (7 x 8 cm).
- Equilibrate the membrane in transfer buffer (Towbin buffer, cat. no. 42558). By use of PVDF membranes equilibrate the membrane first for 2 minutes in methanol and then for additional 5 minutes in transfer buffer.
- Wet the porous pads as well as the four pieces Whatmann 3 MM paper with transfer buffer.
- Remove the gel from the cassette and equilibrate the gel for 5 minutes in transfer buffer.
- Mount the transfer sandwich according to the manufacture’s protocol and place it in the tank blotter.
- Transfer is done at room temperature at 250 mA resp. ca. 60 V for ca. 1 - 2 hours.
After blotting proteins can be stained on the membrane:
- Detection with Ponceau S solution (0.2 %, cat. no. 33427)
- Staining with Amido black