2. Western blotting is a protein detection technique in which mixtures of proteins
are first resolved in a polyacrylamide gel in the presence of sodium dodecyl
sulphate, so that the protein are separated according to molecular weight.
Following electrophoresis, the proteins are transferred and bound to a
microporous membrane, and identified using two parameters:
• Molecular weight, determined by comparing migration to a ladder of proteins of known
molecular weight
• Presence of a signal caused by binding to a specific detection antibody
Because this technique involves multiple steps, and because there is no one set
of optimal conditions applicable to all samples, all target proteins, and all
antibodies, Western blotting researchers often spend countless hours
systematically varying conditions and troubleshooting blots to get the best
signal-to-noise ratios. Nevertheless, Western blots are still very frequently used.
