Western blot (also known as protein immunoblot), a technique used to detect specific proteins in a sample of tissue homogenate or extract, matters much in cell and molecular biology, as well as immunogenetics. It is the most widely used method for detecting protein properties such as qualitative and quantitative detection of tissue antigens, quality of peptide molecules, and detection of antibodies or viral antigens.
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Western blot steps:
Tissue Preparation: Samples are taken from whole tissue or cell cultures.
Gel electrophoresis: The sample protein is separated using gel electrophoresis, which solves the problem of antibody cross-reactivity.
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Transfer: The separated proteins are transferred from the inside of the gel to the membrane.
Blocking: Steps must be taken to prevent interactions between the membrane and the antibodies used to detect the target protein.
Incubation: During this step, the reporting enzyme bound to the modified antibody triggers a colorimetric reaction and produces color when exposed to the appropriate substrate.
Detection: Western Blot is ready to detect labeled probes.
Three key elements for getting to the west point:
• Separation of protein mixtures according to size using gel electrophoresis
• Effective transfer of secreted protein into a strong carrier
• Detection of target proteins with appropriate antibodies
Because western blots can be obtained quickly with simple equipment and cheap reagents, it is one of the most common laboratory techniques. The results obtained are also easy to interpret, unique and unambiguous.