EDC Impressum & DSE

Electrophoresis
Development & Consulting
Dr. Hanspeter Schickle
Wolfgang Gstrein

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Equilibration of the IPG Strip

EppiKlein

IPG-Strip Equilibration Buffer

EquiRehy+LidKlein

Equilibrator-trenches, left side: 12 x 3 ml

Mai 30, 2018

IPG-Strip Equilibration

Weigh in the Urea and the DTT*, res. the IAA** in 2 test tubes (marked with “DTT” and “IAA”),
then add the 6 ml IPG-Strip Equilibration Buffer. Shake till the Urea has been solved (~10 min).
Then add the solutions in 2 tubes (marked with “DTT” and “IAA”) long enough to take the IPG-strips.

Insert the IPG-strip(s) in the Equilibrator, gel side looking up, then add 3 ml of the first (DTT) equilibrating-solutions and shake horizontally each step for 15 min. Repeat this for the second (IAA) equilibrating solution.
See picture above.
* Dithiothreitol  **Iodoacetamide

Step

 

Urea

Thiourea

DTT*

IAA**

Equilibr. Buffer

Time

1

per strip

2.2 g

915 mg

60 mg

---

6 ml (makes ~7.2 ml)

15 min (shake gently)

2

per strip

2.2 g

915 mg

---

150 mg

6 ml (makes ~7.2 ml)

13 min (shake gently)

3

 

---

---

---

---

6 ml gel-buffer

2 min (shake gently)

Normally used:
1   6 M Urea, 2% SDS, 30% glycerol, 50 mM Tris-Cl pH 8.8, 1% DTT*, 0.01% Bromophenolblue, 15 min
2   6 M Urea, 2% SDS, 30% glycerol, 50 mM Tris-Cl pH 8.8, 2.5% IAA**, 0.01% Bromophenolblue, 15 min