Western blotting is a well-established analytical technique for detecting, analyzing, and quantifying proteins. This method is widely used to detect specific protein molecules in complex samples such as tissue homogenates and cell lysates. Western blotting typically involves protein separation by gel electrophoresis followed by transfer to a polyvinylidene difluoride (PVDF) or nitrocellulose membrane. After proteins have been transferred, they can be stained for visualization and directly identified by N-terminal sequencing, mass spectrometry or immunodetection.
In Western blotting immunodetection, proteins are identified through their binding to specific antibodies. Typically, a primary antibody is used in combination with an HRP- or AP-conjugated secondary antibody for chemiluminescent or colorimetric detection using an appropriate substrate. Alternatively, a fluorescently labeled primary or secondary antibody can be used for direct visualization
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