Bio-Rad Mini-PROTEAN® TGX™ Precast Gels for 2-D Electrophoresis - Manual (Page 40)

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Mini-PROTEAN Precast Gels
Running buffer (1x) Dilute 100 ml 10x stock (catalog #161-0744) with 900 ml diH
2
O.
Sample buffer (2x) 200 mM Tris-HCl, pH 6.8, 2% SDS, 40% glycerol, 0.04% Coomassie
(catalog #161-0739) Brilliant Blue G-250, 2% ß-mercaptoethanol or 350 mM DTT (added fresh)
Prepare Buffers
Prepare and Load Samples
Prepare Gels and
Assemble Electrophoresis Cell
Perform Electrophoresis
Remove the comb and tape from the gels and assemble the electrophoresis cell.
Fill the inner and outer buffer chambers with running buffer.
Peptide Analysis (Mini-PROTEAN Tris-Tricine Gels)
Component Reducing Nonreducing
Sample 5 μl 5 μl
Sample buffer
(catalog #161-0739) 4.75 μl 5 μl
β-Mercaptoethanol 0.25 μl —
Total volume 10 μl 10 μl
Heat samples at 90–100°C for 5 min or at 70°C for 10 min.
Load the appropriate amount of sample on the gel.
Connect the electrophoresis cell to the power supply and perform electrophoresis according to
the conditions in the table.
Table A.3. Running conditions for one (1) Mini-PROTEAN Tricine gel in the Mini-PROTEAN Tetra cell . Run conditions
and times are approximate and assume a constant voltage of 100 V. When running more than one gel, current will differ.
16 .5% Gels 10–20% Gels
Power conditions 100 V constant 100 V constant
Expected current (per gel)
Initial 65 mA 65 mA
Final 35 mA 35 mA
Run time 100 min 100 min