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Western blot technique: Definition, principle, theory, procedure, importance
Introduction of the Western Blot Technique, principle, theories, significance
A western blot is a laboratory method used to detect particular protein molecules from between a mixture of proteins. Western blots can also be used to estimate the size of a protein of interest, and to hold the amount of protein expression.
The first step in a western blot is to prepare the protein sample by mixing it with a detergent called sodium dodecyl sulfate, which makes the proteins unfold into linear chains and coats than with a negative charge.
The proteins are transferred from the gel onto a blotting membrane. Although this step is what gives the technique the name "western blotting," the term is typically used to describe the entire procedure.
Once the transfer is complete, the membrane carries all of the protein bands basically on the gel.
The membrane is then incubated with an antibody called the primary antibody, which specifically binds to the protein of interest. Next incubation, any unbound primary antibody is washed away, and the membrane is incubated yet again, but this time with a secondary antibody that specifically recognizes and binds to the primary antibody.
The secondary antibody is linked to a reporter enzyme that produces color or light, which allows it to be quickly detected and imaged. These actions allow a particular protein to be detected from among a mix-up of proteins.
