Immunoblottanalüüs

Immunoblottanalüüs, often referred to as Western blot, is a widely used laboratory technique for detecting specific proteins in a sample. It involves a series of steps to separate, transfer, and identify target proteins. First, proteins are separated by size using gel electrophoresis. This process involves applying an electric current to a gel matrix, causing proteins to migrate at different rates depending on their molecular weight. Following separation, the proteins are transferred from the gel to a solid membrane, typically made of nitrocellulose or PVDF. This transfer is crucial for subsequent detection.

The membrane is then blocked to prevent non-specific binding of antibodies. Primary antibodies, which are specific to the protein of interest, are added to the membrane. These antibodies bind to the target protein. After washing away unbound primary antibodies, a secondary antibody is applied. This secondary antibody is conjugated to an enzyme or a fluorescent molecule and recognizes the primary antibody. Finally, a detection reagent is added, which reacts with the enzyme or fluorescent tag to produce a detectable signal, such as light or color, allowing for the visualization and quantification of the specific protein. This technique is invaluable in molecular biology, immunology, and disease diagnostics for confirming protein expression and studying protein modifications.