Molecular Biotechnology

, Volume 55, Issue 3, pp 217-226

First online:

Open Access This content is freely available online to anyone, anywhere at any time.

A Defined Methodology for Reliable Quantification of Western Blot Data

  • Sean C. TaylorAffiliated withBio-Rad Laboratories (Canada) Ltd Email author 
  • , Thomas BerkelmanAffiliated withBio-Rad Laboratories
  • , Geetha YadavAffiliated withBio-Rad Laboratories
  • , Matt HammondAffiliated withBio-Rad Laboratories


Chemiluminescent western blotting has been in common practice for over three decades, but its use as a quantitative method for measuring the relative expression of the target proteins is still debatable. This is mainly due to the various steps, techniques, reagents, and detection methods that are used to obtain the associated data. In order to have confidence in densitometric data from western blots, researchers should be able to demonstrate statistically significant fold differences in protein expression. This entails a necessary evolution of the procedures, controls, and the analysis methods. We describe a methodology to obtain reliable quantitative data from chemiluminescent western blots using standardization procedures coupled with the updated reagents and detection methods.


Western blot Densitometry Protein expression