C.B.S. Scientific EBU-204 - Manual (Page 11)

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SECTION 4
Applications & Running Conditions for Tank Type Electro-Blotting
4.1
Recommended Buffers, Membranes, Power Settings and Transfer Times
Gel Type Blotting Buffer Transfer Membrane Power
denaturing: SDS-
PAGE
(Western)
ClearPAGE Transfer Buffer
cat. # FB82500 as follows:
for 1:20 dilution use 50ml
ClearPAGE Transfer buffer,
200 ml Methanol and 770ml
Ultrapure water
(“Towbin”: 25mM
Tris/192mM Glycine/ pH
8.3,
Methanol 20% (w/v)
nitrocellulose/
nylon PVDF: ion-
exchange
· room temperature, 70V-100V,
200mA, 1.5- 4 hours
· 4ºC, 180V, @160mA, 45
minutes - 2 hours
· Overnight, 30V, 75mA
non-denaturing:
SDS-PAGE (acidic
or neutral proteins)
25mMTris/192mM Glycine,
pH 8.3
25mM sodium phosphate,
pH6.5
15mM sodium borate,
pH9.2
nitrocellulose;
nylon PVDF; ion-
exchange
diazo-paper
diazo-paper
· room temperature, 60V,
200mA, 4 hours
· 4ºC, 125V, 400mA, 1.5 hours
·
Overnight, 30V, 100mA
IEF (native-basic
proteins, acid urea
gels)
0.7% acetic acid (w/v)* nitrocellulose; nylon
diazo-paper
· room temperature, 70V,
400mA, 4 hours
· 4ºC, 100V, 600mA, 1.5 hours
· Overnight, 30V, 200mA
DNA
(Southern)
1XTAE or agarose
0.5X TBE, acrylamide
nylon; +charged
nylon, neutral or
+charged
· room temperature, 30-40V,
25-150mA, 2-4 hours
· 4ºC, 125V, 400mA, 1.5 hours
RNA
(Northern)
19.6mM phosphate/5.4mM
citrate
pH3.0 or 25mM sodium
phosphate
pH 6.5
nylon nitrocellulose · room temperature, 60V,
200mA, 4 hours
· 4ºC, 125V, 400mA, 1.5 hours
· Overnight, 30V, 200mA