The production of antibodies is dependent on the humoral response of living cells to the entry of foreign antigens by injection. A powerful response is obtained following the injection of foreign molecules, cells or viruses, but some substances are not very immunogenic.
Either antigen or host can be subjected to alterations to enhance the immune response. Until now, the nature of such manipulations was discovered by trial and error. However, current knowledge of how the immune system works has enabled the discovery of better methods.
Many naturally occurring and synthetic compounds act as potent immunogens, including proteins, carbohydrates, peptides, lipids, nucleic acids, but in different ways. For instance, non-protein molecules and peptides must be conjugated to a protein carrier, such as bovine serum albumin or hemocyanin from the keyhole limpet, to make them immunogenic. The carrier protein provides the binding site for class II T receptor binding sites required for immunogenicity, which would otherwise be lacking due to the small size of the antigen.
Another additional step is the co-administration of an adjuvant to improve the quantity or quality of the immune response. An adjuvant is a substance that stimulates the immune response in a nonspecific manner. The use of an adjuvant enables the reduction of the antigen quantity required to obtain a prolonged antibody response.
Figure 1. Hybridoma production
For polyclonal antibodies to be produced, an antigen needs to be injected repeatedly over a long period, with several weeks intervening between successive doses. This stimulates the antibody producing B cells that are specific for that antigen to switch to large-scale antibody production.
The host animal’s blood now contains a wide range of antibodies against multiple epitopes present on the antigen. This immune serum can be used as such to obtain large quantities of several specific antibodies to the same antigen. Alternatively, it may be subjected to affinity purification to isolate the different components of the serum, and obtain specific antibodies.
In order to produce monoclonal antibodies, the procedure is followed as above up to the point of repeated immunization. Subsequently, all the B cells are filtered out and fused with immortal cells from tumor cell lines to form hybridomas, each of which is capable of churning out a specific antibody to a particular epitope forever. These are then tested to assess the level of antibody production and the practical advantage of the antibody.
Hybridomas responsible for producing a particular antibody are named using clone names. These are purposely assigned on a unique basis so that they can be identified by them. They are then cloned using isolation techniques and cultured using tissue culture methods. Another technique of producing antibodies is to clone genes that code for specific antibodies into transfection vectors, leading to the production of recombinant antibodies.
The difference between polyclonal and monoclonal antibodies is that the latter are identical to each other, reacting specifically to a single epitope. The hybridoma or recombinant cell secretes the antibody into the culture medium, from where it can be harvested and used as such or separated into its purer components by the use of affinity chromatography.
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