Bio-Rad GS-900™ Calibrated Densitometer - Manual (Page 47)

Step Voltage, V Ramp Time Units
1 8,000 Rapid 26,000 Vh
2 750 Hold
1 250 Rapid 0:20 HH:MMr
2 8,000 Gradual 1:00 HH:MM
3 8,000 Rapid 26,000 Vh
4 1,500 Hold
1 8,000 Rapid 20,000 Vh
2 750 Hold
1 250 Rapid 0:20 HH:MMr
2 8,000 Gradual 1:00 HH:MM
3 8,000 Rapid 20,000 Vh
4 750 Hold
1 250 Rapid 0:20 HH:MMr
2 8,000 Gradual 1:00 HH:MM
3 8,000 Rapid 32,000 Vh
4 750 Hold
1 250 Rapid 0:20 HH:MMr
2 8,000 Gradual 1:00 HH:MM
3 8,000 Rapid 40,000 Vh
4 750 Hold
1 8,000 Rapid 29,000 Vh
2 750 Hold
1 250 Rapid 0:20 HH:MMr
2 8,000 Gradual 1:00 HH:MM
3 8,000 Rapid 29,000 Vh
4 750 Hold
Table 9.3. Preprogrammed protocols for 11 cm ReadyStrip IPG strips.
Protocol Name
11 cm pH 5.5–6.7
11 cm pH 6.3–8.3
11 cm pH 7–10 R
11 cm pH 7–10 G
11 cm pH 3–10 R
11 cm pH 3–10 NL R
11 cm pH 4–7 R
11 cm pH 5–8 R
11 cm pH 3–10 G
11 cm pH 3–10 NL G
11 cm pH 4–7 G
11 cm pH 5–8 G
11 cm pH 3–6 R
11 cm pH 3.9–5.1
11 cm pH 4.7–5.9
11 cm pH 3–6 G
R = rapid, G = gradual
Step Voltage, V Ramp Time Units
1 4,000 Rapid 15,000 Vh
2 500 Hold
1 250 Rapid 0:20 HH:MMr
2 4,000 Gradual 1:00 HH:MM
3 4,000 Rapid 15,000 Vh
4 500 Hold
1 4,000 Rapid 10,000 Vh
2 500 Hold
1 250 Rapid 0:15 HH:MMr
2 4,000 Gradual 1:00 HH:MM
3 4,000 Rapid 20,000 Vh
4 500 Hold
1 250 Rapid 0:15 HH:MMr
2 4,000 Gradual 1:00 HH:MM
3 4,000 Rapid 20,000 Vh
4 500 Hold
1 250 Rapid 0:15 HH:MMr
2 4,000 Gradual 1:00 HH:MM
3 4,000 Rapid 25,000 Vh
4 500 Hold
1 4,000 Rapid 16,000 Vh
2 500 Hold
1 250 Rapid 0:15 HH:MMr
2 4,000 Gradual 1:00 HH:MM
3 4,000 Rapid 16,000 Vh
4 500 Hold
Table 9.2. Preprogrammed protocols for 7 cm ReadyStrip ™ IPG strips.
7 cm pH 3–10 R
7 cm pH 3–10 NL R
7 cm pH 4–7 R
7 cm pH 5–8 R
Protocol Name
7 cm pH 5.5–6.7
7 cm pH 6.3–8.3
7 cm pH 7–10 R
7 cm pH 7–10 G
7 cm pH 3–10 G
7 cm pH 3–10 NL G
7 cm pH 4–7 G
7 cm pH 5–8 G
7 cm pH 3–6 R
7 cm pH 3–6 G
7 cm pH 3.9–5.1
7 cm pH 4.7–5.9
R = rapid, G = gradual
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2-D Electrophoresis Guide Methods Chapter 9: First-Dimension IEF with IPG Strips
The protocols and settings described are for IEF using
the PROTEAN i12 IEF cell. Preprogrammed protocols
serve as convenient starting points for optimization of
IEF conditions (Tables 9.2 to 9.5).
■ ■
The recommended focusing temperature for most
samples is 20°C
■ ■
For better sample entry, start IEF with a low voltage
gradient (200 V for 30–180 min) and limit current to
50 µA per IPG strip for the whole run
■ ■
Focusing time depends on gel length, pH gradient,
gel additives, and protein amount loaded.
Vertical streaking is often caused by overfocusing —
isoelectric precipitation (pI fallout) increases with
focusing time. For this reason, do not conduct first-
dimension IEF for any longer than is necessary
■ ■
After completion of the IEF run, IPG strips should
be stored frozen at –80°C in rehydration trays
or immediately applied to a second-dimension
SDS-gel. Frozen IPG strips can be stored for
about 3–6 months
IEF Programming Recommendations