User Manual

Section 5
Appendix
A Buffer and Reagent Preparation
To avoid introducing experimental error when using this kit, we suggest using premixed buffers
and stains. These can be found in section 6, Product Information.
1X Tris/glycine/SDS running buffer (25 mM Tris, 192 mM glycine, 0.1% SDS, pH 8.3)
Tris base 3.03 gm
Glycine 14.4 gm
SDS 1.0 gm
Water to 1 L, do not adjust pH
Coomassie Brilliant Blue R-250 stain solution (0.1% Coomassie Blue R-250 in 40% MeOH, 10%
HOAc)
Coomassie Blue R-250 1.0 gm
Water 500 ml
Acetic acid 100 ml
Methanol 400 ml
Destain solution (10% acetic acid, 40% methanol)
Water 500 ml
Acetic acid 100 ml
Methanol 400 ml
B Programming the PROTEAN IEF Cell
New method programming instructions are also found in the PROTEAN IEF cell manual.
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1. - Select the NEW METHOD mode to program a new method
REHYDRATION >
PRESET METHOD >
STORED METHOD >
NEW METHOD >
2. - Use the alphanumeric keypad to enter a program name.
- Select NO for Rehydration
- Use default FOCUS TEMP of 20°C
- Press NEXT
NAME: "Ecoli" >
REHYDRATION Yes NO >
FOCUS TEMP 20 °C >
NEXT >
3. - Enter step 1 final voltage of 250 V
same voltage for all strip lengths)
- Select LINEAR ramp. Use key next to arrow, >, to move
the brackets, [ ] ,around the selection
- Press NEXT
S 01 250>
SELECT VOLTAGE SLOPE
LINEAR [
ää
]>
NEXT >