Western Blotting Principle
The fundamental principle of western blotting: separating proteins by size, transferring them to a membrane, and detecting a target protein with antibodies.
Overview
Western blotting (immunoblotting) is a technique used to detect specific proteins in a sample. The principle rests on three main stages: separation of proteins by molecular weight, transfer of those proteins onto a solid support, and selective detection using antibodies.
Core idea
Proteins are first separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) according to size. They are then transferred (blotted) onto a membrane, where they can be probed with a primary antibody that binds the protein of interest, followed by a labeled secondary antibody or other detection method to visualize the band.
Why it works
The combination of size-based separation and antibody-based detection gives high specificity and allows estimation of molecular weight and relative abundance. Content here can be enriched with diagrams and more detail on SDS-PAGE and transfer conditions.