There are four main methods of detection in Western blotting: radioisotopes, chromogenic substances, chemiluminescence and fluorescence. Radioisotopes and chromogenic substances were first used to detect proteins bound to a membrane. These earlier ...

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Western blotting is an extremely sensitive method. The sensitivity and success of the technique is directly dependent on the antibodies that are used for detecting the protein of interest. Primary antibodies directly bind to the protein of interest, ...

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Membranes used for Western blotting have a high affinity for proteins allowing binding and retention of transferred proteins. Before transferred proteins can be detected using antibodies, the membrane must be incubated with a blocking buffer to ...

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Efficient transfer of proteins out of a gel onto a membrane is critical when performing a Western blot. Inefficient transfer of a protein may skew results or cause the protein to become undetectable on the blot. High molecular weight proteins are ...

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