2 dna and rna blotting membrane – Bio-Rad Trans-Blot® Plus Cell User Manual

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PVDF Membrane
Bio-Rad offers PVDF (Polyvinylidene difluoride) membranes that are ideal for
immunoassays of blotted proteins (Immun-Blot PVDF) or amino-terminal sequencing
and amino acid analysis (Sequi-Blot PVDF). PVDF retains proteins under extreme
conditions, such as exposure to organic solvents or acidic or basic conditions.
Greater protein binding capacity allows for better retention of easily transferred
proteins, while allowing more time or higher voltages to transfer difficult or larger
proteins. Greater protein retention during sequencing manipulations enhances the
likelihood of obtaining information from rare, low abundance proteins, by increased
initial coupling and more consistent yields. In addition, PVDF membrane exhibits
better binding efficiency of blotted material in the presence of SDS in the transfer
buffer. PVDF must first be wetted in 100% methanol but can then be used in a
transfer buffer that does not contain alcohol.

Nitrocellulose Membrane
Nitrocellulose membranes have been used extensively for protein binding and
detection

2,6,8-10

. Nonspecific protein binding sites are easily and rapidly blocked on

nitrocellulose, avoiding subsequent background problems. No pre-activation of the
membrane is required. With nitrocellulose, low molecular weight proteins (especially
those <20,000 Da) may be lost during post transfer washes, thus limiting detection
sensitivity

. Smaller pore size nitrocellulose membrane (0.2 µm), has been shown

to be effective in eliminating this loss

. Large proteins (those >100,000 Da) that

are denatured by SDS may transfer poorly to nitrocellulose if alcohol is added to the
transfer buffer. Alcohol in the transfer buffer increases binding of SDS-proteins to
nitrocellulose, but decreases pore sizes in the gel.

5.2 DNA and RNA Blotting Membrane

Zeta-Probe

Nylon Membrane

Zeta-Probe membrane is an ideal alternative to nitrocellulose for the analysis of
nucleic acids. The membranes bind nucleic acids in low ionic-strength buffers,
making electrophoretic transfer of nucleic acids from agarose and acrylamide gels
possible. Zeta-Probe membrane allows efficient binding of all sizes of single
stranded DNA and RNA in the presence of low ionic strength buffers

. Unlike

nitrocellulose, Zeta-Probe membranes can be hybridized as many as 20 consecutive
times.