User Manual

Section 4
Troubleshooting Guide
Problem Cause Solution
1 Smile effect – band a. Center of the gel running a. Improper cooling. Ensure
pattern curves upward at hotter than either end buffer level is to the fill
both sides of the gel line indicated on the tank.
Use cooling coil and
refrigerated circulator to
maintain buffer temperature
of 20 ºC
b. Power conditions are b.Decrease voltage from 200 V
excessive to 150 V.
2. Run takes unusually a. Ion depletion in running a. Prepare and use fresh
long time. buffer (lower tank) buffer.
b. Running buffer too b. Check buffer protocol
concentrated. and dilute buffer if
necessary.
3. Changes in protein a. Ion depletion in running a. Prepare and use fresh
mobility or band buffer (lower tank) buffer.
sharpness.
4. SDS is precipitating. a. Running buffer is too cold. a. Set the refrigerated
circulator to maintain a
buffer temperature of
18–20 ºC.
5. Vertical streaking of a. Sample overload or a. Dilute sample,
proteins. protein precipitation selectively remove
predominant protein in the
sample, or reduce the
voltage by about 25% to
minimize streaking.
b.The ratio of SDS to protein
should be enough to coat
each protein molecule,
generally 1.4:1. It may
require more SDS for some
membrane samples.
c. Ensure sample is suspend-
ed completely in sample
buffer prior to loading.
6. Lateral band spreading a. Diffusion out of the a. Minimize the time
wells prior to turning between sample
on the current application and power
start up.
b. Ionic strength of sample b. Use same buffer in sample
lower than that of gel as in gel or stacking gel
7. Skewed or distorted a. Salts in sample a. Remove salts by
bands dialysis, desalting column,
etc.
8. Lanes constricted at the a. Ionic strength of sample a. Desalt sample and
bottom of the gel. higher than that of neighboring samples.
surrounding gel.
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