It is common to see references to the “specificity” or “greater specificity” of monoclonal antibodies compared to polyclonal antibodies. Is that claim justified? No one would dispute the fact that a monoclonal is directed toward a single site or epitope of the target protein, while a polyclonal may contain antibodies directed toward multiple epitopes on the protein. However, in my opinion, that fact alone does not make monoclonal antibodies more specific. Indeed the single epitope toward which a monoclonal is directed may in fact be shared by many different proteins in addition to the protein of interest. Such a monoclonal would not be specific even though it recognizes only a single epitope. In contrast, a polyclonal antibody raised against the same protein may contain antibodies directed toward that same non-specific epitope as the monoclonal, as well as other epitopes that are more specific. In such a situation the serum of the polyclonal would at least contain some antibodies that are specific and thus it would be “more specific” than the monoclonal. Moreover, it may be possible using affinity purification to isolate the specific antibodies. This may seem to some as a trivial issue but it can be extremely important in IHC where it is quite difficult to control for cross reactivity. Thus simply opting for a monoclonal is no guarantee of specificity and one must still utilize a full range of specificity controls. An example of such a non-specific monoclonal is shown in the figure 1.  Note that the monoclonal antibody recognizes the NET protein at ~50 kDa but it also recognizes proteins at 75 and 95 kDa.

 1447-NET

Figure 1. The monoclonal raised against the NET protein labels three 
prominent bands at 50, 75 and 95 kD in a lysate of rat cortex.

One additional issue relating to this question is the difference in the purification/selection of monoclonal and polyclonal antibodies. The process of purification and selection of monoclonal antibodies rarely involves screening for specificity.  Thus the hybridoma screens of hundreds or thousands of clones typically rely solely on the ability to recognize a target in a plate assay.  There is typically no selection for affinity for the target and or specificity. This contrasts with polyclonal antibody affinity purification which, as its name suggests, can preferentially yield higher affinity and higher specificity antibodies. 
Taken together these comments are not at all meant to minimize the very real importance of monoclonal antibodies in clinical environments. Rather these comments are meant to underscore the importance of specificity controls in using all antibodies and to show that, in some cases, polyclonal antibodies may be more specific than monoclonal antibodies.

Summary
Are Monoclonal Antibodies Really More Specific?
Article Name
Are Monoclonal Antibodies Really More Specific?
Description
Epitopes can be shared between many proteins, so a monoclonal antibody that only recognizes one epitope may not be more specific than a polyclonal antibody that recognizes multiple epitopes on the same protein.
Author
Mike Browning
Publisher Name
PhosphoSolutions
Publisher Logo

3 thoughts on “Are Monoclonal Antibodies Really More Specific?

  1. Interesting blog article. It should be noted that with respect to the NET protein there are also two glycosylation sites which can cause additional bands. The core protein will run ~55 kD and the gycosylated protein can be detected at ~75-90 kD (all seen in your figure above). It is important that end user fully understand their protein. Other bands can be detected due to phosphorylation, unique cleavage sites, protein degradation, incomplete protein processing, improper sample handling… In these cases, the antibody is recognizing the many different forms of the protein. It would be interesting to see how a NET pab stained the same blot.

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