The Protein Man's Blog | A Discussion of Protein Research

Recombinant Antibodies: An Overview

Posted by The Protein Man on Oct 28, 2015 10:00:00 AM
The Protein Man

Recombinant antibodies (rAbs) are monoclonal antibodies which are generated in vitro using synthetic genes. Unlike monoclonal antibodies (mAbs) which are produced using traditional hybridoma-based technologies, rAbs do not need hybridomas and animals in the production process.

Both monoclonal and recombinant antibodies can be used in biomedical science and toxicological research, and are effective therapeutic treatments for cancer, autoimmune disorders and a host of other diseases. However, while monoclonal antibodies have become one of the most common tools in biomedical science and medicine due to their ability to bind and neutralize or destroy cell-specific antigens, the ascites method of production causes significant pain and discomfort to the animals used in the process.

As such, the governments of Australia, Germany, the Netherlands and the United Kingdon banned it in favor of in vitro methods. The US also endorses the use of in vitro methods as the default procedure for the production of mAbs.

However, it is important to note that in vitro methods involving hybridomas also have their own limitations, which include the following:

  • Require immunization and subsequent euthanasia of the animals used in the process.
  • Slow and labor-intensive.
  • Often cause immune reactions so the antibodies need to be altered and "humanized" before they can be administered to patients.

Production of Recombinant Antibodies

Basically, recombinant antibodies are monoclonal antibodies generated in vitro using synthetic genes. The technology involves recovering antibody genes from source cells, amplifying and cloning the genes into an appropriate phage vector, introducing the vector into a host (bacteria, yeast, or mammalian cell lines), and achieving expression of adequate amounts of functional antibody.

Recombinant antibodies can be cloned from any species of antibody-producing animal provided there are appropriate oligonucleotide primers or hybridization probes available. The ability to manipulate the antibody genes also makes it possible to generate new antibodies and antibody fragments (Fab fragments and scFv) in vitro. Display libraries, commonly expressed in phage or yeast, can then be analyzed to select for desirable characteristics arising from such changes in antibody sequence.

How do you select the ones that display your desired antibody? You can do it through a process called panning. One of the simplest panning procedures is a variation of common ELISA procedure. To do this, you incubate the antibody library with the target immobilized on a solid. Any unbound phage is removed through washing and specifically bound phages are eluted and amplified by infecting E. coli cells. The process is repeated 3 to 4 times to isolate the phages that are displaying antibodies with the highest affinity and stability. Genes for the selected antibodies are sequenced and subjected to affinity maturation. The genes for the best antibodies can then be transferred into an appropriate expression system for large-scale production.

The Advantages of Using Recombinant Antibodies

  • Increased reproducibility and control. While researchers often lose control of the antigen once they have injected it into the host animal in hybridoma-based systems, recombinant antibody production allows them more control over the antigen. Since rAbs are defined by the sequences that encode them, they are more reliable and provide more reproducible results than mAbs. By adjusting experimental conditions, researchers can easily favor the isolation of antibodies against antigens or antigen characteristics.
  • Decreased production time. Using recombinant antibody technology, an antigen-specific antibody can be produced in as little as eight weeks. On the other hand, hybridoma technology requires a minimum of four months to produce a suitable antibody.
  • Isotype conversion. A desirable recombinant antibody fragment can easily be converted into a different species, isotype or subtype by adding the appropriate constant domain. This makes it easier to switch antibodies into a more preferable format.
  • Animal-free technology. Recombinant antibodies can be produced without using any animals in the process. This eliminates the numerous ethical and animal welfare concerns commonly associated with traditional monoclonal antibody production.

Topics: Antibody Production

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