Protein Protocols Protein Protocols

Discontinuous buffer systems
Tris-glycine buffer with addition of SDS Cathodic side: 8.3 The presence of 20% methanol at the anodic side facilitates 17
at the cathodic side and methanol at the 0,1% (w/v) SDS the dissociation of protein–SDS complexes and the
anodic side (Svoboda buffer) 25 mM Tris Glycine proteinbinding to the negative charge matrix, while
192 mM the presence of 0.1% SDS at the cathodic side increases
(no methanol) the rate of transfer of cationic and/or hydrophobic at the
cathodic side increases the rate of transfer of cationic
and/or hydrophobic proteins.
Anodic side:
25 mM Tris 8.3
192 mM Glycine
20% (v/v) Methanol
Kyhse–Andersen buffer Cathodic side: This system uses isotachophoresis to elute proteins from 18
40 mM 6-amino-n-hexanoic acid 9.4 from the gel. SDS, glycine, and SDS–protein complexes
25 mM Tris from the gel move toward the anode according to
20% (v/v) Methanol their net mobilities in the stated order. At the cathodic
side, 6-amino-n-hexanoic acid is the terminating ion
Anodic side: and thereby, carries away the proteins from the
300 mM Tris 10.4 polyacrylamide gel.
20% (v/v)
Methanol
(in contact with the anode)
25 mM Tris 10.4
20% (v/v) Methanol
(in contact with the membrane)
Laurière buffer Cathodic side: This system of buffers introduces a difference of pH 19
0.4% (w/v) SDS 8.4 which remains stable during the entire electro-
60 mM lactic acid transfer time period and an asymetrical
100 mM Tris disposition of methanol and SDS on each side
(in contact with the cathode) of the gel–membrane pair.
(continued)
Semidry Protein Blotting 325