Polyclonal Antibodies

Polyclonal antibodies (pAbs) are a mixture of different antibodies that all recognize the same protein. They are produced by immunizing an animal, usually a rabbit, goat, sheep or donkey, with the target of interest, allowing for its recognition by circulating B lymphocytes. When a B lymphocyte binds to its target antigen via the unique antibody displayed at the B cell surface, it undergoes clonal expansion to form a pool of identical plasma cells that all produce the same antibody. To harvest the antibody, blood is collected from the host animal and the serum extracted. The serum can then either be used directly, or the antibody may be purified, typically using Protein A, Protein G, or an immobilized form of the antigen. Because any protein immunogen will have multiple epitopes, each of which is recognized by a different B cell clone, and because the resultant antibody preparation is the product of multiple B cell clones, it is described as being polyclonal. An advantage of polyclonal antibodies is that they can improve assay sensitivity as a result of multiple antibody molecules binding to different epitopes on the same protein of interest. This can be especially useful when detecting scarce targets, or proteins that are likely to change conformation, as well as when performing techniques such as immunohistochemistry (IHC), immunoprecipitation (IP), or chromatin immunoprecipitation (ChIP), when certain epitopes may be masked by fixation, crosslinking, or the formation of protein complexes. The ability of polyclonal antibodies to recognize multiple epitopes can also give them broader application functionality compared to monoclonal antibodies. For example, a monoclonal antibody that is validated for IHC-based detection of a native protein might not be able to recognize the same (denatured) protein by western blotting (WB), yet a polyclonal antibody might work well in both applications. Other advantages of polyclonal antibodies include their relatively low cost, faster availability when a new target is discovered, and (in the case of rabbit polyclonal antibodies) better success rates when raised against more challenging antigens. On the flipside, a limitation of polyclonal antibodies is that they are only available in finite supply, corresponding to the lifespan of the host. In addition, the manner in which polyclonal antibodies are produced means that they can exhibit batch-to-batch variability, both between different lots collected from the same animal over time and between different animals. Polyclonal antibodies may also cause a higher level of background noise than monoclonal antibodies as they are more likely to cross-react with other proteins sharing similar epitopes to the target. It is common to address the risk of non-specific binding by performing cross-adsorption, such as by cross-adsorbing secondary antibodies against species other than that against which they were raised. Critically, by designing experiments carefully, researchers can mitigate non-specific binding to have greater confidence in their data.