Medical Sciences 4900F/G Lecture Notes - Lecture 4: Western Blot, Primary And Secondary Antibodies, Polyvinylidene Fluoride
22 Apr 2019
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Document Summary
Last week you loaded your protein samples and ran them on an sds-page. Once the proteins were separated on the gel you transferred the proteins to a pvdf membrane and dried it out. This week you will be probing the membrane with antibodies so that you can visualize the protein of interest. Antibodies have a specific affinity for a particular part of an antigen the epitope. The antibody that binds to the target protein is called the primary antibody (1 ab). Ab is localized using a secondary antibody (2 ab: secondary antibody is used for th indirect detection of target antigens. The 2 ab is conjugated to a reported enzyme that detects and visualizes the antibody. Its important that the secondary antibody used is directed against iggs from the species of origin of the primary antibody iggs = one of the five antibody isotypes iggs provide the majority of antibody-based immunity against invading pathogens.
