6.3 Incorrect Band Size

Troubleshooting

1. Inaccurate Indication by Pre-Stained Protein Marker
The migration characteristics of pre-stained protein markers may change when electrophoresed under buffer conditions with different molecular weights, which can lead to certain deviations. If precise localization of protein molecular weight is required, an un-stained protein marker can be used.
2. Excessive Cell Passage Number
Thaw and use cells with a low passage number for the experiment.
3. Inherent Differences Between Cells/Tissues of Different Species
Judgments on whether a band is the target band should be based on the results of overexpression, knockout/knockdown experiments.
4. Protein Modifications
Proteins with different modifications exhibit different molecular weights. Even for the same type of modification, there are variations in the degree (e.g., high glycosylation vs. low glycosylation). To determine the specific modification type, relevant literature should be consulted or gel excision followed by mass spectrometry should be performed.
5. Protein Multimers
Ensure sufficient denaturation of proteins by adding an adequate amount of reducing agent and boiling the protein sample.