Monoclonal versus polyclonal antibodies – it is a widespread issue in research laboratories and biopharmaceutical campaigns.
In this article, we want to outline the basics on the definitions of the terms “monoclonal” and “polyclonal” and then move on to discuss their differences and similarities with information regarding their respective sources, functionality, production concepts, pros & cons and example applications. Moreover, we will give some insight into what distinguishes modern recombinant antibodies from traditional monoclonal antibodies.
Polyclonal antibodies (pAbs) are by definition antibodies that are produced by a population of genetically different B cells in response to being exposed to an antigen (immunization). Each single antibody is specific to a distinct part of the antigen’s surface, an epitope. Therefore, polyclonal antibodies are a relatively heterogeneous set of antibody molecules.
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Polyclonal antibodies are made in B cells of animals (e. g. rabbits, mice, horses and humans) after being exposed to an antigen that elicits an adequate immune response. The resulting immunoglobulins (such as IgG) can be extracted from a blood donation after removing cellular blood constituents and clotting proteins.
Read more: Antibody production by B cells: 7 facts
The vast majority of pAbs is manufactured from laboratory animals, but antibody extracts from convalescent human donors can be used as antisera against toxins and pathogens (this was done in the early phase of the COVID-19 pandemic). Off-target effects and background reactivity can be reduced by subjecting the polyclonal antibodies to affinity purification.
Polyclonal antibodies come with the big advantage of a relatively small price tag. They are comparably easy to produce without animal lab infrastructure and have shorter lead times of about 100 days.
Moreover, they excel in detecting low concentrations of target protein due to their binding of several different epitopes on an antigen and boast very quick binding kinetics. Labeling usually does not interfere with specificity.
Disadvantageous is the dependency on laboratory animals. Each animal has an individual immune system and expresses more or less significantly different antibodies, leading to reproducibility issues.
So when would you use a polyclonal antibody? Polyclonals are the antibody of choice when single-epitope specificity is not a high priority and cost-efficiency is required. While the latter is a given, single-epitope specificity is often overpowered and actually hinders detection at low levels. A single antigen might be bound by numerous labeled polyclonal antibodies, thus amplifying the signal, while monoclonal antibodies bind at a 1:1 ratio at the single epitope. Hence, pAbs are predestined for diagnostic applications with low detection limits.
While monoclonal antibodies (mAbs) are historically defined as antibodies that are derived from a single clone of B type immune cells, nowadays the term also encompasses recombinant antibodies manufactured in mammalian cell cultures with very high molecular homogeneity and single-epitope specificity just like mAbs. Examples are modern therapeutic recombinant antibodies which are colloquially called monoclonal antibodies even though they were never near a B cell during the production process.
But how are monoclonal antibodies produced in the lab? Traditionally, monoclonal antibody production was based on the fusion of single B lymphocytes with immortal myeloma cells (a type of cancer cell) to create antibody-producing and immortal cell lines: hybridoma cells. Those cells are either transplanted into host animals or grown in culture. This was the first process capable of producing large quantities of mostly homogeneous antibodies.
With the recombinant revolution of biology the engineering of peptides, enzymes, cytokines and even antibodies at the genetic level became possible. This type of antibody is called recombinant antibody and it is produced in vitro in a suitable expression system, preferably in mammalian cell lines such as HEK and CHO cells.
Probably the biggest advantage of monoclonal antibodies (mAbs) is the possibility to produce very homogeneous antibodies on scale, especially when recombinant technology and cell culture processes are used. This is the prerequisite for antibody therapy: a steady supply of reproducibly high quality drug compound.
Additionally, monoclonal antibodies show low cross-reactivity and low background noise which is required in sensitive biochemistry experiments or diagnostic applications.
A significant disadvantage of mAbs is the price tag and usual lead time. That being said, evitria as an innovative antibody expression service provider offers project finishing times from sequence to antibody as short as 5 weeks. Traditional production times are often in the 6 month range.
When would you use monoclonal antibodies – whenever therapeutic antibodies are the drug of choice: cancer, immunology, and anti-infectives are the biggest fields. The possibility to optimize human antibodies at the DNA sequence level and express them with intact post-translational modifications allows very high biologic specificity leading to high efficacy with low adverse effects.
This is a typical example of the high-level functionally being more important than the price tag. Another one is immunoassays when high specificity (e. g. among closely related target antigens) is required. Monoclonals are ideally suited as primary antibodies to capture the antigen of interest.
Since antibodies are products of the vertebrate immune system, their traditional sources are animals. This fact has led to numerous public ethical discussions whether their use in research and medicine is worth the sacrificing of laboratory animals.
The harnessing of recombinant technology has allowed scientists to overcome this dilemma by developing methods using in vitro laboratory techniques and cell culture to manufacture recombinant monoclonal antibodies. The mammalian cell lines were derived from a single tissue sample and are able to multiply on their own: the CHO cell line stems from a Chinese Hamster ovary cell. Hence, high quality antibodies are routinely produced without any laboratory animals.
Polyclonal and monoclonal antibodies have a set of major differences, influencing the way they are produced and work. This, on the other hand, has a great impact on the fields of applications they are eligible for, and the advantages and disadvantages they come with.
Here is an overview of the main differences between polyclonal and monoclonal antibodies:
Despite several similarities, monoclonal and polyclonal antibodies can already be distinguished by their respective definition. These differences primarily depend on where they stem from, and what their main features consist of.
polyclonal Abs monoclonal Abs definition by source primary host animal secondary host animal after hybridoma graft transplantation, hybridoma cell culture, recombinant mammalian cell culture by functionality high variability in binding sites on antigens specific to a single epitope on an antigen
Monoclonal and polyclonal antibodies have major differences considering their production. This also has an impact on ethical considerations on these types of antibodies.
Production of polyclonal Abs Production of monoclonal Abs purification of blood from immunized host animals ascites liquid from hybridoma-transplanted host animals, hybridoma cell culture, recombinant mammalian cell culture
Both monoclonal and polyclonal antibodies have a set of convincing advantages. Nevertheless, both antibody types also come with considerable downsides, as illustrated in the table below:
polyclonal Abs monoclonal antibodies + high sensitivity for specific antigen high specificity for single epitope + relatively quick and easy high homogeneity + low background, low cross-reactivity – cross-reactivity relatively long lead time – batch-to-batch variability batch-to-batch variability
Based on their main characteristics, features and production, monoclonal and polyclonal antibodies have several, often different fields of application. Nevertheless, both are highly relevant tools in life sciences and medicine.
Applications of polyclonal Abs Applications of monoclonal Abs sandwich elisa therapeutic antibodies immunoprecipitation antibody drug conjugates immunohistochemistry highly selective immunoassays Western blotting very specific reagents immunofluorescence assay