Hoefer TE 22, User Manual, Page: 27 / 34

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p19
Protein transfer buffers
Use a buffer with low ionic strength, such as the
two listed below, to prevent overheating. Use the
alternate CAPS buffer when Tris cannot be used,
as in peptide sequencing. CAPS can improve
transfer because of its effect on the charge of
the protein (see Matsudaira, 1987). For native
proteins, we suggest using the electrophoresis
buffer for transfer as well. Use the Towbin
buffer to transfer SDS-denatured proteins
toward the anode.
Towbin buffer
(25 mM Tris, 192 mM glycine, 20% v/v methanol,
pH 8.3, 2 liters)
Tris (FW 121.1) 25 mM 6.0 g
Glycine (FW 75.07) 192 mM 28.8 g
SDS a (FW 288.4) 0.1% (3.5 mM) 2.0 g
Dissolve in 1.5 liters distilled water. Add methanol as
required
b
. Bring to 2 liters with distilled water. Do not
adjust the pH, which should be between 8.2 and 8.4.
Optional: Chill before use.
a Optional: Adding SDS can improve transfer efficiency.
b Depending on the membrane type selected, adding methanol can
improve the transfer results (see discussion and table above). Because
buffers containing methanol may deteriorate if stored for long periods,
add methanol as required just prior to transfer.
CAPS buffer, 1X
(10 mM CAPS, pH 11.0, 2 liters)
CAPS (FW 221.3) 10 mM 4.44 g
[3-(cyclohexylamino)-1-propanesulfonic acid]
Dissolve in 1.5 liters distilled water, adjust to pH 11.0 with
conc. NaOH. Adjust volume to 2.0 liters.