All tags Antibody guide A comparison between polyclonal and monoclonal

A comparison between polyclonal and monoclonal

Key differences, advantages and disadvantages of polyclonal and monoclonal antibodies.

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Summary

Polyclonal antibodiesMonoclonal antibodies
Inexpensive to produceExpensive to produce
Skills required are lowTraining is required for the technology used
Time scale is shortTime scale is long for hybridomas
Produces large amounts of non-specific antibodiesCan produce large amounts of specific antibodies
Recognizes multiple epitopes on any one antigen Recognizes only one epitope on an antigen
Can have batch-to-batch variabilityOnce a hybridoma is made, it is a constant and renewable source
-No or low batch-to-batch variability

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Polyclonal antibodies

Facts:

  • Recognize multiple epitopes on any one antigen. Serum obtained will contain a heterogenous complex mixture of antibodies of different affinity.
  • Polyclonals are made up mainly of IgG subclass.
  • Peptide immunogens are often used to generate polyclonal antibodies that target unique epitopes, especially for protein families of high homology.
  • Most polyclonals are either goat or rabbit polyclonals. However, other species can be used (e.g. mouse, rat, chicken).

Antibody production:

  • Inexpensive to produce.
  • Technology and skills required for production less complex compared to monoclonal antibodies.
  • Production time scale is short.
  • Polyclonal antibodies are not useful for probing specific domains of antigen because polyclonal antiserum will usually recognize many domains.


Advantages:

Polyclonals will recognize multiple epitopes on any one antigen which has the following advantages:

  • High affinity. Polyclonals can help amplify signal from target protein with low expression level, as the target protein will bind more than one antibody molecule on the multiple epitopes. This would not be an advantage for quantification experiments (e.g. in flow cytometry), as the results would become inaccurate.
  • Due to recognition of multiple epitopes, polyclonals can give better results in IP/ChIP.
  • Polyclonals are more tolerant of minor changes in the antigen (e.g. polymorphism, heterogeneity of glycosylation or slight denaturation) than monoclonal (homogenous) antibodies.
  • Polyclonals will identify proteins of high homology to the immunogen protein or can be used to screen for the target protein in tissue samples from species other than that of the immunogen (e.g. polyclonal antibodies are sometimes used when the nature of the antigen in an untested species is not known). This also makes it important to check immunogen sequence for any cross-reactivity.
  • Polyclonal antibodies are often the preferred choice for detection of denatured proteins.
  • Multiple epitopes generally provide more robust detection.


Disadvantages:

  • Prone to batch-to-batch variability.
  • They produce large amounts of non-specific antibodies which can sometimes give background signal in some applications.
  • Multiple epitopes make it important to check immunogen sequence for any cross-reactivity.
  • Polyclonal antibodies not useful for probing specific domains of antigen, because antiserum will usually recognize many domains.

Monoclonal antibodies

Facts:

  • High specificity. Detect only one epitope on the antigen.
  • They will consist of only one antibody subtype (e.g.  IgG1, IgG2, IgG3). Where a secondary antibody is required for detection, an antibody against the correct subclass should be chosen.
  • Monoclonals are typically rat or mouse monoclonals. However, monoclonals can be generated from various species such as rabbit and goat. 

Learn more about Abcam Rabbit Monoclonal technology and its advantages.


Antibody production:

  • High technology required.
  • Training is required for the technology used.
  • Timescale is long for hybridomas.


Advantages:

  • Once hybridomas are made, it is a constant and renewable source and all batches will be identical - useful for consistency and standardization of experimental procedures and results.

Monoclonals detect one epitope only on any one antigen which has the following advantages:

  • Monoclonals usually have less background from staining of sections and cells. As they are more specifically detecting one target epitope, they are less likely to cross-react with other proteins.
  • Due to their high specificity, monoclonal antibodies are excellent as the primary antibody in an assay, or for detecting antigens in tissues, and will often give significantly less background staining than polyclonal antibodies.
  • Compared to polyclonal antibodies, homogeneity of monoclonal antibodies is very high. If experimental conditions are kept constant, results from monoclonal antibodies will be highly reproducible, between experiments (e.g. very low lot-to-lot variation). 
  • Specificity of monoclonal antibodies makes them extremely efficient for binding of antigen within a mixture of related molecules, such as in the case of affinity purification.


Disadvantages:

  • Monoclonal antibodies may be too specific (e.g. less likely to detect in across a range of species).
  • More vulnerable to the loss of epitope through chemical treatment of the antigen than polyclonal antibodies are. This can be offset by pooling two or more monoclonal antibodies to the same antigen (e.g. cocktail antibodies).

See examples of cocktail antibodies from Abcam.

Abcam's RabMAb®​ technology combines the benefits of both monoclonal and polyclonal technology to create highly specific and senstitive rabbit monoclonal antibodies.

Learn more about the advantages of RabMAb®​ technology.


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