Bio-Rad Model 225 Tube Gel Casting Stand - Manual (Page 16)

2. First-dimension gels do not focus a. Check the tube gels, tubing connectors,
well. sample reservoir and ends of the gels
for bubbles.
b. Check the pH of the upper and lower
chamber buffers. Increase the NaOH
concentration of the buffer in the upper
chamber.
c. Pre-electrophorese the tube gels prior to
loading samples.
d. Increase the focussing time.
e. Reduce the salt concentration in the
sample.
3. Streaking in the second-dimension
gels. (Refer to the Mini-PROTEAN cell
instruction manual for further
troubleshooting.)
Horizontal streaking: a. Sample preparation problems. Add
detergent containing sample buffer,
then centrifuge at 100,000 x g to
remove the insoluble pellet.
b. Overloaded protein sample. Load less
protein.
c. Nucleic acids bound to the protein.
Treat sample with an endonuclease.
d. Focusing time not optimized. Perform a
time course to determine the best
volt-hours for the samples.
Vertical streaking: a. Problems in the sample. Determine if
the vertical streaks are related to the
sample by running an SDS-PAGE
molecular weight standard on the same
gel.
b. Pinpoint streaking in the background of
the gel may be caused by dust or
bacteria in the water supply. Filter all
the solutions through a 0.45 µm
nitrocellulose membrane.
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