Bio-Rad Mini-PROTEAN® TGX™ Precast Gels for 2-D Electrophoresis - Manual (Page 38)

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Mini-PROTEAN Precast Gels
SDS-PAGE (Mini-PROTEAN TGX Gels)
Running buffer (1x) Dilute 100 ml 10x stock (catalog #161-0732) with 900 ml diH
2
O.
Sample buffer Use Laemmli sample buffers: catalog #161-0737 or catalog #161-0747 (4x).
Prepare Buffers
Prepare and Load Samples
Prepare Gels and
Assemble Electrophoresis Cell
Perform Electrophoresis
Remove the comb and tape from the gels, rinse wells, and assemble the electrophoresis cell.
Fill the inner and outer buffer chambers with running buffer.
Component Reducing Nonreducing
Sample 5 μl 5 μl
Laemmli sample buffer, 2x
(catalog #161-0737) 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.1. Running conditions for SDS-PAGE in the Mini-PROTEAN Tetra cell. Standard conditions are constant 300 V.
100 V 200 V 300 V
Run time 85–95 min 30–40 min 15–20 min
Expected current (per gel)
Initial 15–20 mA 25–50 mA 55–75 mA
Final 5–10 mA 20–31 mA 45–70 mA
Expected temperature 25°C 25–35°C 30–45°C
Outer buffer volume
1–2 Gels 2-gel mark 2-gel mark 4-gel mark
3–4 Gels 4-gel mark 4-gel mark 4-gel mark 1
1 Requires the PowerPac ™ HV or PowerPac Universal power supply.