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Polyclonal Antibodies

By Susha Cheriyedath, MSc

Polyclonal antibodies (pAbs) are a complex mixture of several antibodies which are usually produced by different B cell clones of an animal. These antibodies can recognize and bind to many different epitopes of a single antigen and hence can form lattices with the antigens.

Production of Polyclonal Antibodies

PAbs are produced using a series of steps as described below:

Antigen Preparation

The quality of antigen used directly affects the immune response obtained. Even small amounts of impurities will lead to antibodies reacting more to the impurity than to the desired antigen. The quantity of the antigen is also crucial as it is proportional to the immune response evoked. Too little or too much antigen may cause sensitization, suppression, or tolerance. Thus, the antigen quantity should be determined based on several factors such as properties of the particular antigen, the animal species chosen, injection frequency and the purity level of the antigen.

Animal Species Selection

The animal species for the production of pAbs should be selected depending on factors such as:

  • the amount of PAb required,
  • the phylogenetic relationship between the animal and antigen,
  • the age of the animal,
  • the ease of obtaining blood samples, and
  • the application in which the PAb is going to be used.

Commonly used animal species in the lab for pAb production are the rabbit, rat, mouse, guinea pig, hamster, goat, chicken, and sheep. Rabbits are more preferred due to their size and relatively long life span. However, in order to produce larger quantities of pAbs, farm animals such as goats, sheep, and horses are used.

Immunization Protocol

The immunization protocol differs for different animal species and adjuvants used. Adjuvants are prepared compounds routinely used as a form of stimulus in the case where the induced immune response would otherwise be inadequate.
The smallest possible volume of antigen capable of inducing an effective immune response is injected into the animal. The injection route needs to be selected carefully for different animal species. The injection of antigen can be carried out as one single volume or as several low volumes at different sites. Booster injections are provided at regular intervals of weeks or months depending on the antigen used during the first immunization. Antibody titers measured after a booster injection can help to determine if more booster injections are required.

Post-Immunization Observation

Animals are monitored daily to evaluate the side effects of immunization and blood is withdrawn at regular intervals. Serum from the animals can be analyzed to monitor the antibody responses and extract the antibodies.

This is a relatively inexpensive process and it can be used to isolate a large quantity of an antibody in a single extraction.

Applications of Polyclonal Antibodies

Some key applications of PAbs are given below:

  • Pharmaceutical industry:
  • Digoxin Immune Fab is a pAb used to treat the fatal digitoxin toxicity.
  • Rho(D) immune globulin is a pAb used to prevent a maternal immune response that could transfer hemolytic disease to the newborn baby. The antibody binds passively to the antigen and is produced from a pool of human plasma collected from Rhesus-negative donors who have antibodies for the D antigen (present on red blood cells).
  • Histopathological analyses that use immunoperoxide staining
  • Affinity purification of antigens
  • Sandwich ELISA technique to detect tumor markers

Advantages of Polyclonal Antibodies

Some advantages of pAbs are listed below:

  • PAbs are a heterogeneous mix of antibodies that can bind to wide range of antigenic epitopes. Hence, small change in the epitopes of an antigen is less likely to affect pAbs.
  • PAbs are very stable across a wide range of salt concentrations and pH values.
  • PAbs have a higher specificity compared to monoclonal antibodies (mAbs) as they have specific antibodies for each epitope of an antigen.
  • PAbs are relatively easy to manufacture in a timely and cost-efficient way.

Disadvantages of Polyclonal Antibodies

Some disadvantages of pAbs are listed below:

  • Unlike mAbs, pAbs produced against the same antigen in different animals will be different in the case of immunized animals.
  • The affinity of pAbs to antigens may change over time.
  • The amount of pAbs produced is limited by the size and lifespan of the animal.
  • The purity and concentration levels of a specific antibody are lower in pAbs than that in mAbs.

References

Last Updated: Jun 9, 2016

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