5 Important Factors in the Production of Antibodies

by Martin Gray Content Writer

Antibody production is a process that involves multiple steps and precise attention to certain factors that are critical for an antibody that works.

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Here is a summary of the five key factors in producing an antibody: it works or selects the commercial antibody that best suits clinical tests.

Key factors for antibody production:

1. Selection and obtaining the antigen

It is essential to analyze the target against which we want to obtain the antibody:

Type of antigen (protein, peptide, small molecules, carbohydrates)
Size
Availability of sufficient quantity for immunizations and validations or the possibility of obtaining it
Homology with other proteins that you do not want to detect

2. Enhance the immunogenicity of the antigen

The antigen must elicit an immune response strong enough in the animal to obtain the antibodies of interest. Immunogenicity can be increased by:

Carriers or large molecules (KLH, BSA) attached to small antigens (peptides)
Adjuvants that act as catalysts for the immune response, increasing its magnitude. These adjuvants make it possible to increase the antibody titer in a relatively short period. The most common is Freund's adjuvant but on certain occasions, other alternatives such as aluminum salts can be used.

3. Selection of the host or animal in which the antibodies will be produced

This selection will determine the type of antibody to be obtained:

Rabbits: They produce an intense and very fast immune response, which is why they are usually the species of choice for obtaining polyclonal antibodies.
Mice: It is one species that generate the best performance of immune cells for the development of hybridomas, so it is usually the species of choice for obtaining monoclonal antibodies.
Goat: Like rabbits, the immune response is intense, with the addition that, as it is a larger animal, the volume of antibody that can be obtained is higher.
Camels/Llamas: They produce what are known as "Single domain Antibodies," significantly smaller in size than IgG, with greater ease to bind to small molecules.
Hens: They produce a special type of immunoglobulins, IgY, and obtain antibodies against highly conserved mammalian proteins.

4. Type of antibody to be obtained

Polyclonal antibodies: The antibodies will be titrated during the immunization process to ensure that the final bleeding is done at the highest titer time and thus obtain the highest amount of antibody possible. It must be taken into account that the immune response experiences different dynamics in different animals, so the moment in which the titer peak occurs may vary from one species to another.
Monoclonal antibodies: once the immune response has developed, the animal's spleen cells will be extracted from which the hybridomas will be obtained. The production of antibodies from these can be done in cell culture (in vitro) or ascites fluid (in vivo).

5. Test in which the antibody will be used

The same antibody can work perfectly in one technique and give a diffuse signal in another. We remind you of these tips to optimize your ELISA, Western Blot, and Immunohistochemistry assays.