Monoclonal or Polyclonal Antibody, which is right for Me?

Monoclonal or Polyclonal Antibody, which is right for Me?

By: Stephen Pelsue PhD, Science Director, Antibody Services

One of the most common questions I get when discussing antibody development is: which is better a  monoclonal or polyclonal antibody? The simple answer is, it depends on the application. For example, if  specificity is the driving factor then a monoclonal antibody likely is your best bet; however, if you are looking to  use the same antibody source for several applications, a polyclonal may best serve your needs.

Before we dive into the advantages or disadvantages of a monoclonal antibody or polyclonal antibodies, let’s first step  back and think about the composition of each one. As many antigens are structurally complex (proteins, bacteria, virus,  etc.), when used as an immunogen they will elicit a broad response and many B cell clones will produce antibodies  specific to the antigen. A polyclonal antibody is a “pool” of several antibodies that recognize distinct epitopes on the same  antigen. A monoclonal antibody is, as the name indicates, an antibody that is produced by a single clone, or more  specifically derived from a single B cell and therefore, the antibody recognizes a single target structure or epitope.

While many comparisons of monoclonal versus polyclonal antibodies and antibody services focus on the differences in  cost and time, I would like to focus on the functional differences between monoclonal and polyclonal antibodies. Let’s go  through a few of the critical variables that are important for immunoassays and discuss the benefits and weaknesses for  both monoclonal and polyclonal antibodies.

Specificity

Specificity, in the context of antibody binding, refers to its ability to uniquely distinguish an antigen from other components that might be present. This may refer to the recognition of a circulating protein in the bloodstream from all the other compounds in the blood, or the ability to bind to a specific virus or bacterial strain, while not recognizing closely related species or variants. A monoclonal antibody would recognize a single epitope on the antigen, while polyclonal antibodies would recognize many different epitopes on the specific antigen. Therefore, the monoclonal antibody only detects or binds to the antigen if its single epitope is available, while the polyclonal antibody can recognize several epitopes and bind to the antigen even if not all epitopes are available. Which of these properties best suits your needs may depend on the level of specificity required and the amount of cross-reactivity with similar proteins or strains.

This requires us to consider the whole antigen as a compilation of all its epitopes. Let’s consider a soluble monomeric protein that is 40kD and does not contain any repetitive sequences or regions. For the sake of discussion let’s say there are 20 potential epitopes, of which 5 are found on structurally similar proteins (cross-reactive epitopes), and 2 of the epitopes contain sequence variation (variant epitopes) within the sample population (due to disease mutations, evolutionary variation, strain variation, etc.). If the monoclonal and polyclonal antibodies recognized only the epitopes which were unique non-cross-reactive epitopes and not the variant epitopes, there would be little to no difference in the specificity between them. However, if the polyclonal recognized multiple epitopes that were unique as well as the 5 epitopes that were cross-reactive with structurally similar proteins, then the polyclonal would no longer be specific to the desired protein of interest. The specificity of a monoclonal, however, would be defined by which individual epitope it recognized. In the case of a variant epitope, its ability to be recognized would be defined by the epitope, for example if a monoclonal recognizes only one of the variants then it will be very specific for that individual variant. Conversely a polyclonal could contain antibodies that recognize the same epitope but different variants, and therefore the polyclonal could recognize all variants.

This same principle could extend from an individual protein to a bacterial cell or virus, which would be far more complex, as there are many more possible epitopes. Getting back to the question of “which is better”, would depend on the goal for the particular immunoassay of interest. If cross-reactivity is not a concern, then either would likely work well. If specificity is the major defining requirement a monoclonal would likely be better as the individual antibody that binds to the appropriate unique epitope could be selected and provide increased specificity over a polyclonal pool of antibodies. However, if all variants, species, or protein family is of interest in being captured, then an individual epitope may not allow for this goal to be met and would require a polyclonal antibody, either by using polyclonal sera (or purified IgG from sera) or a mix of multiple monoclonal antibodies to achieve this goal. So, it is critical to define the goals of the assay and what the specificity needs are to fully determine if a monoclonal antibody (or pair) versus a polyclonal antibody pool would be the appropriate choice.

Use in Applications

While monoclonal and polyclonal antibodies can be used in all the same assay formats and platforms, there are several aspects to consider which approach may be best for an individual use. How an antigen or epitope is presented will make a difference in a specific antibodies performance in a given assay. An antibody that work magnificently in one assay may not perform well at all in a different platform. This can be due to antibody-antigen binding kinetics or also the availability of the epitope(s). For example, a conformationally dependent epitope will likely not be appropriate for a Western Blot or other assay that denatures the protein prior to antibody binding.

The great benefit to monoclonal antibodies is that their targeting of a single epitope allows them to be selected for very specific capabilities and then the assay can be optimized to provide reproducible performance. However, as they are typically developed and selected with a specific purpose in mind, changing platforms or conditions will likely require a new antibody if the conditions are substantially different from the original use. This is where polyclonals can be of great use as they contain several antibodies that recognize several epitopes, some linear, some conformational and providing much more coverage of the antigen. This will allow for broader usage in multiple immunoassay platforms.

The other consideration is the management of supply. As polyclonal antibodies are generally generated in larger animals (rabbits, goats, sheep, etc.) they can be immunized and maintained for long periods of time. The difficulty is that the “blend” of antibodies will shift or drift over time, so that the antibodies collected from one production bleed to another may require optimization each time in an assay. To get around this, large quantities can be produced and purified (as appropriate) and used as a single lot over a period of time. However, if a new lot of antibody needs to be produced it may not be similar to the original lot even if the same approach is used. As monoclonal antibodies are generated from a cell line, they can be reproducibly manufactured to be able to generate equivalent antibody lots year in and year out.

Considerations

The point of this discussion is to highlight the fact that the best approach is dependent on the goals and purpose of the immunoassay. Monoclonal antibodies and polyclonal antibodies have their benefits and weaknesses but understanding the target and the immunoassay platform will allow the “best” approach to be defined. One must also consider the lifespan of its use, supply chain, and manufacturing requirements as well to be able to make a clear choice.

If you would like to discuss whether a monoclonal or polyclonal antibody development is best suited to your assay, 
please contact our technical team at sales@mainebiotechnology.com or Int: +(1) 207-797-5454

Learn more about our Monoclonal Antibody Development here or Polyclonal Antibody Development here



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