"The method used in molecular biology to detect specific proteins. This technique involves the preparation of samples with a mixture of proteins, electrophoresis, gel protein transfer from gel to membrane, incubation with appropriate antibodies and detection of the desired protein."
- Western blot definition
Western blot has found its application in the diagnostics of diseases, e. g. in the case of hepatitis C, this method detects the virus core. It is also used for diagnosis of borreliosis and tuberculosis. In HIV diagnosis, a positive Western blot test performed after a positive ELISA test gives 99.9% confidence. Western blot technique is useful in assessing the quality of protein purification and recombinant protein production after gene cloning.
Immunobloting is used in the diagnosis of bacterial diseases, such as borreliosis caused by Borelia burgdorferi Tuberculosis caused by Mycobacterium tuberculosis. This technique can also be used to diagnose viruses such as HIV, SARS-induced coronavirus and herpes flushing virus HSV, which is an etiological agent of herpes. Western blot can also be used to detect some parasitic diseases, such as anthrosis caused by tapeworm larvae and even genetic diseases, such as certain forms of Fanconi anaemia. Literary data indicates that it is also possible to use this technique in cancer diagnostics, which proves its usefulness and versatility.
Western blot technology is used to detect and identify proteins. The procedure consists of several parts. The first step is the separation of the protein mixture in the polyacrylamide gel. The proteins are then transferred to the membrane (in our case it is semi-dry) which does not specifically bind all proteins. The transfer takes place in the direction of the positive electrode. After electrophoretic separation in the presence of SDS (ionic, negatively charged, detergent) the proteins are denatured and all have a negative charge proportional to their mass. All of them will then move towards the positive electrode. Therefore, the membrane should be laid in such a way that it is placed on the path of migration of proteins from the gel. There are two types of transfer: wet transfer and semi-dry transfer.
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