EDC Impressum & DSE

Electrophoresis
Development & Consulting
Dr. Hanspeter Schickle
Wolfgang Gstrein

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SDS-Electrophoresis Trouble Shooting

Mai 30, 2018

General mistakes
When mounting up a horizontal electrophoresis run, several mistakes can be done.
Normally their occur already after starting by watching the power supply and the migration of the dyes:

No currant, 0 mA, no migration of the dyes:
1   The cables inside the chamber are not switched properly
2   The platin electrodes are not in contact with the electrode strips
3   The electrode strips are not in contact with the gel (no overlapping zone)
4   The power supply is not programmed in a correct way (no volt = no mA)

 Too fast migration of the dyes, gel is sweating and later burning:
The dyes of the samples and the gel should migrate around 0.5 mm per minute, not more!
1   Too much currant, too high mA value:
     a   Power supply is not programmed correctly
     b   A part of the gel runs with the conditions of a whole gel
2   The cooling device is not running correctly, or it is not pumping really through the cooling plate

 Gel is leaving its support film:
Do not use staining recipes with alcohol concentrations higher than 40%.
The gel will shrink or swell to much so the forces between the film and the gel get to high!

Troubles-MeOH

Samples
Even in a excellent performing electrophoretic system you will not get  automatically acceptable results from every sample!
There will be always 'trouble-making-samples', mostly coming from:

[complex sample mix]
[fat in the samples]
[salt in the samples]
[protein concentration]
[samples striking]
[electro-dekantation]
[run too short]
[no alkylation]
[basic proteins]

1) Complex sample mix (protein & fat) form a channel and no bands occur

2) Bands in a channel and smearing (too much fat)

3) Bands unsharp, tend to go to the neighbour-lanes (too high salt concentration)

4) Bands curved (too high protein-concentration)

5) Bands are striking, not uniform (Glycoprotein or very lipophylic proteins)

6) Sample disturbs the whole run. Anodal slot-edge is burning

7) Run is too short, separation pattern is not  reaching the anode

8) Sample is changing by storage-time. Big proteins appear at the starting point

9) Bands unsharp also in low concentration, smearing. Unsufficient separaton

1,2) Striking track, too much fat

3) Too much salt/buffer in the sample

complex mix

complex sample, too much fat

fat in samples
salt in samples

slightly too much salt:
bands are not as sharp as the standard-proteins

Too much salt / buffer

far too much salt:
lanes go to the neighbour-lanes

4) Too high sample concentration

Sample too high concentrated

4) The edges of a series of samples occur curved

TSSerieKlein

5) Bands not uniform

NRun

6) Slot is burning

Electro Dekantation

7) Run is too short

8) Sample is changing

9) Bands unsharp smearing also in low concentration

run is oo short
Reoxidation 2
Reoxidation 2
basic proteins
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