Bio-Rad Immun-Blot® PVDF Membrane - Manual (Page 5)

tion. At this point, the membrane is ready
to bind proteins in any blotting application.
3. After the membrane has been wetted with
buffer, do not allow it to dry (white spots
will form where the membrane is dry).
Protein will not bind to the dried mem-
brane, and dry spots will not rewet in aque-
ous solutions. If the membrane becomes
dry prior to blotting, repeat steps 1 and 2 to
rewet it.
Section 3
Dot Blotting
Dot blotting requires special precautions to
insure that protein is bound to the PVDF mem-
brane before it dries. Directly spot protein to the
wetted membrane and allow it to dry. Vacuum
assisted dot or slot blotting is not recommend-
ed, due to the potential for the PVDF to dry out
before the proteins are bound.
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transfer buffer. After equilibration, the mem-
brane can be used in a semi-dry, tank, or capil-
lary blotting system with any acidic or basic
blotting buffer.
Note: Always handle membranes using gloves or
forceps to prevent contamination.
1. Immerse the membrane in 100% methanol
for a few seconds, until the entire mem-
brane is translucent. In methanol, it wets
immediately. (Solutions containing 50%
methanol concentration can be used to pre-
wet the membrane.)
2. Transfer the wetted membrane to a vessel
containing transfer buffer or water.
Incubate in buffer until it is equilibrated
(² 2–3 minutes). The membrane will float
on the surface of the buffer until complete-
ly equilibrated. After it is equilibrated it can
be easily submerged into the aqueous solu-
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