6.4 Others

Troubleshooting

1. White Bands on Dark Background
Possible Causes
Excessive sample loading volume
Excessively high concentrations of primary antibody and secondary antibody
Solutions
It is recommended to reduce the protein loading volume.
Decrease the concentrations of the antibodies.
2. "Smiling" Bands (Bands Curving Upward at Both Ends)
Possible Causes
Inhomogeneous gel
Excessively high electrophoresis temperature and overly fast migration rate
Aging of the electrophoresis apparatus
Solutions
It is recommended to reduce the voltage and current for electrophoresis, and reconfigure the gel.
Replace with a new electrophoresis apparatus.
3. Diffuse Bands or Band Tailing
Possible Causes
Excessive protein loading volume
Protein degradation
Excessive cell debris
Prolonged delay after electrophoresis stops
Nucleic acid contamination
Solutions
It is recommended to reduce the protein loading volume.
Avoid repeated freezing and thawing of samples.
Centrifuge the sample before loading.
Perform protein transfer as soon as possible after electrophoresis is completed.
Add ultrasonic treatment or use needle aspiration (to disrupt samples) as an alternative.
4. "Dumbbell-Shaped" Bands (Hollow in the Middle)
Possible Causes
Gel preparation issues: When removing the comb, the bottom of the wells protrudes, causing proteins to settle toward both ends.
Impurities in the sample push the protein toward both ends.
Solutions
It is recommended to pull the comb slowly and vertically upward to avoid deformation of the gel wells.
Protein impurities can be removed by centrifugation.