Western Blot Protocols-Antibody Incubation & Gel Visualization

Western Blot Protocols-Antibody Incubation & Gel Visualization Following the blocking step, the protein of interest can be detected using antibodies.

This following process are very similar to ELISA. Many tips and cautions in ELISA are equally applied in western blot. Both monoclonal and polyclonal antibodies can be used for Western blotting analysis (Table 2). The most two important standard while choosing an antibody are: 1) weather it can recognize the denatured proteins; and 2) weather it can cause cross reaction. Polyclonal antibodies tend to be more sensitive, but are less specific than monoclonal antibodies. Monoclonal antibodies, on the other hand, tend to be more specific but less sensitive. Polyclonal antibodies are usually chosen for their relatively lower price and less time consuming to produce. • Primary antibodies should be raised in species as distinct as possible from the sample species: it is better to raise a primary antibody against a mouse protein in a rabbit, for example, rather than a rat.