Western Blot (Immunoblot)

Analytical technique separating proteins by gel electrophoresis and detecting specific targets on a membrane with antibodies.

Specifications & Metrics

type: Protein Immunodetection
readout: Chemiluminescent, fluorescent, or colorimetric band intensity at a specific molecular weight
measures: Protein expression levels, post-translational modifications (phosphorylation, cleavage), and antibody specificity
format: SDS-PAGE gel electrophoresis followed by electroblotting onto nitrocellulose or PVDF membrane
pioneer: Invented in 1979 by Harry Towbin et al. and Jaime Renart et al.; named in 1981 by W. Neal Burnette

Independently invented in 1979 by Harry Towbin, Theophil Staehelin, and Julian Gordon and by Jaime Renart, Jakob Reiser, and George Stark, the method was named "Western blot" by W. Neal Burnette in 1981 as a play on Edwin Southern's DNA blot.
Proteins are denatured in sodium dodecyl sulfate (SDS) to impart a uniform negative charge-to-mass ratio, separated by size through polyacrylamide gel electrophoresis (SDS-PAGE), and transferred onto a nitrocellulose or PVDF membrane.
Phospho-specific primary antibodies allow pharmacologists to track pathway inhibition (such as phospho-ERK1/2 after U0126 treatment or phospho-S6K after rapamycin) alongside total protein loading controls.