Hoefer SE900 - Manual (Page 28)

• p22
Problem Solution
Instrument Problems:
No voltage or current at start of the run
Instrument not properly attached to power supply. Make sure high voltage
leads are connected into correct +/- terminals and is secure. In some cases
adapters may be required.
Broken electrode. Check continuity of wire with a volt meter.
Lid not secured properly onto banana plugs. Reposition and check lid is fit
securely in place.
Break in HV lead line. Check continuity of wire with a volt meter.
2D Results Problems:
Vertical streaking of sample down from the
IPG Strip not properly placed on gel surface. Avoid gouging separating
top of the gel towards the bottom of the gel gel while loading strips.
Perform iodoacetamide treatment.
Make sure IPG strip uniformly contacts the gel surface along the entire
length of the strip.
Overloading of protein.
Horizontal streaking of proteins Poor sample preparation.
Inadequate focusing.
Poor contact between the IPG strip and the second dimension gel.
Spots are skewed or distorted Gels run too fast-uneven migration.
Overlay the resolving gel with water-saturated n-butanol before polymeriza-
tion begins to avoid forming an uneven gel surface.
Uneven gel polymerization or gradient formation. See casting problems for
more support.
IPG strip not properly placed on gel surface. Avoid gouging separating gel
while loading strips.
Sample entry into the second dimension gel is run at too high a power
setting. Run gels at recommended run conditions.
Bubbles present in second dimension gel will distort the spot migration.
No proteins visible on second dimension Quantity loaded too little for detection method. Increase protein load
gel after staining or try a more sensitive staining method.
Too little sample applied to first dimension gel. Check protein concentration
of sample.
Equilibration steps too short or too long. Perform each equilibration step
for 10–15 minutes.
Blank regions in the second dimension gel Tris, salts, and SDS can cause alterations in the focusing position of proteins
in the first dimension. Reduce or eliminate these compounds from the
first dimension.
Bubbles between the strip and the second dimension gel.