For more information on Western blot normalization, watch these webinars:
- Western Blot Normalization: What You Need to Know
- Normalization Methods for Quantitative Western Blot Analysis
Using a total protein stain to detect the total protein in each lane of your gel or blot is becoming more popular. Total protein staining is a direct measure of the total amount of sample protein in each lane. For each lane, the sum of all the signal intensities of all the proteins in the lane is used for normalization.
This more direct approach may increase the accuracy of normalization. Unlike housekeeping proteins, total protein staining does not require validation for each experimental context.
A total protein stain should produce a linear increase in signal intensity in response to increasing protein concentration. It should also correct for variation at all points in the Western blot process, including gel loading and transfer to membrane. It must be compatible with downstream immunodetection of your blot. You should make sure that the signal intensity of the total protein stain is moderate, without saturation or low signal-to-noise ratios.
REVERT™ Total Protein Stain provides linear, proportional signal across a broad range of sample concentrations.
Learn more about total protein controls in the full paper on normalization: Western Blot Normalization: Challenges and Considerations for Quantitative Analysis