Bio-Rad Immun-Blot® PVDF Membrane - Manual (Page 6)

SDS imparts a net negative charge to the
proteins and helps facilitate movement out
of the gel. Binding of proteins to Immun-
Blot PVDF membrane is not affected by the
addition of this amount of SDS.
3. PVDF should be clean and free of wrinkles.
4. Wet the membrane following the protocol
in the membrane wetting section.
5. Make sure there are no bubbles between the
membrane and the gel.
6. After transfer, rinse the membrane three
times (5 minutes each) with distilled water.
Solutions
Towbin buffer:
25 mM Tris 3.03 g
192 M glycine 14.4 g
20% methanol 200 ml
Adjust volume to 1 liter with dd H
2
O.
Prechill the buffer before use.
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Section 4
Electrophoretic Blotting
Immun-Blot PVDF membrane can be used
with a variety of transfer equipment, including
Bio-Rad’s Trans-Blot cell, Mini Trans-Blot
cell, and Trans-Blot SD Semi Dry cell. In gen-
eral, tank blotting is more quantitative with
higher binding yields than semi dry transfers.
However, very good results can be obtained
when a semi dry apparatus is used with a well
optimized gel and transfer system. The semi
dry transfer is much faster, with transfer times
as low as 15 minutes versus tank blotting
which requires 1 to 3 hours for transfers.
1. For most proteins, use a Towbin buffer 1
with methanol (MeOH).
2. For proteins that resist transfer out of the
gel, up to 0.1% SDS can be added to the
transfer buffer. This stabilizes the effect of
methanol in stripping SDS off the proteins.
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