Secondary antibodies are antibodies that recognise and bind the constant region of other antibodies. These are used in any application that uses indirect methods of detection for proteins such as Western blot, ELISA, immunoprecipitation, immunofluorescence, immunohistochemistry, flow cytometry. Secondary antibodies are designed to recognise the constant regions of specific species of antibodies, thus allowing multiplexing and detection of many different target proteins in tandem.
Typically, secondary antibodies are pre-adsorbed to prevent cross-reactivity with antibodies of other species. This is often done by passing the secondary antibody solution through a column that contains immunoglobulins from other, non-target species. Secondary antibodies that cross-react with other species of antibodies are unable to elute and are thus removed from the final solution. Most distributors and manufacturers employ this quality-control, which helps prevent non-specific binding in experiments and thus improves reproducibility of results.