{"id":2267,"date":"2026-05-20T12:26:20","date_gmt":"2026-05-20T10:26:20","guid":{"rendered":"https:\/\/bioeducator.eu\/?page_id=2267"},"modified":"2026-05-20T12:31:05","modified_gmt":"2026-05-20T10:31:05","slug":"western-blot","status":"publish","type":"page","link":"https:\/\/bioeducator.eu\/?page_id=2267&lang=en","title":{"rendered":"Western blot"},"content":{"rendered":"\n

Of all the immunochemical techniques, the Western blot is a unique method. It is the only one that uses electrophoretic<\/a> separation as a starting point for analysis. Although it is time-consuming, it has found widespread use in laboratories around the world.<\/p>\n\n\n\n

\"Western<\/figure>\n\n\n\n

What is a Western blot?<\/h2>\n\n\n\n

Western blotting is a multi-step analytical technique. It is used to detect specific proteins in biological samples, such as cell lysates, tissue homogenates and body fluids.<\/p>\n\n\n\n

How does a Western blot work?<\/h3>\n\n\n\n

A Western blot analysis can be divided into several stages:<\/p>\n\n\n\n

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  1. Protein preparation:<\/strong> This stage may vary depending on the matrix being analysed. In the case of cells, lysis must be performed to extract the proteins. Sometimes it is necessary to isolate proteins from cell membranes using detergents. Some matrices may require the use of various techniques, such as dialysis, ultrafiltration, SPE, etc.<\/li>\n\n\n\n
  2. SDS-PAGE:<\/strong> The isolated proteins are separated by electrophoresis. Their separation is based on molecular weight. You can find out more about SDS-PAGE in this article<\/a>.<\/li>\n\n\n\n
  3. Transfer to a membrane:<\/strong> proteins trapped in the pores of the gel are difficult to analyse further. The gel is fragile and impermeable to antibodies. It is therefore necessary to transfer the proteins from the gel to a membrane.<\/li>\n\n\n\n
  4. Blocking the membrane<\/strong>: As with the ELISA technique, the membrane must be blocked. This prevents non-specific binding of antibodies to the membrane itself. BSA or milk proteins are most commonly used for blocking.<\/li>\n\n\n\n
  5. Immunodetection: <\/strong>Labelled antibodies are used to detect proteins. These bind specifically to the target protein. Any excess antibodies must be washed away. Depending on the detection method chosen, the proteins appear as coloured or fluorescent bands.<\/li>\n<\/ol>\n\n\n\n
    \"The<\/figure>\n\n\n\n

    Where does the name \u2018Western Blot\u2019 come from?<\/h2>\n\n\n\n

    The history of the naming of blotting techniques is one of the most fascinating stories in molecular biology.<\/p>\n\n\n\n

    It all began in 1975, when Edwin Southern<\/strong> described a method for transferring DNA fragments onto a membrane. The technique was named the Southern blot<\/strong>, after its inventor.<\/p>\n\n\n\n

    Two years later, a similar method was developed for RNA; the scientists, playing on the analogy with the cardinal directions, called it the Northern blot<\/strong>.<\/p>\n\n\n\n

    In 1979, Harry Towbin and his colleagues described a method for transferring proteins, but it was W. Neal Burnette who, in 1981, coined the term „Western blot<\/strong>„. Burnette, who was working at the Fred Hutchinson Cancer Research Centre at the time, wanted to continue the tradition started by Southern and the „northern” variant for RNA. Although the editors of scientific journals initially rejected the name as too colloquial, the scientific community embraced it with enthusiasm<\/p>\n\n\n\n

    Types of protein transfer<\/h2>\n\n\n\n

    Transfer\u2014the process of transferring proteins from the gel to the membrane\u2014is a critical step that determines the sensitivity and reproducibility of the entire experiment. The proteins must retain the pattern (known as the electrophoretic map) established in the gel.<\/p>\n\n\n\n

    Wet transfer<\/h3>\n\n\n\n

    This is the most traditional and efficient method. The gel and membrane are sandwiched between two plates and completely immersed in a tank containing the transfer buffer.<\/p>\n\n\n\n