Antigen selection

If you contact us for the generation of a new antibody, several aspects have to be focussed on before a new project is initiated. Some of the most important factors are choice and design of the antigen as well as the screening strategy employed to identify and select the best antibody-secreting hybridoma. We assist in antigen design and in choosing the best strategy and we implement the project. Close communication is maintained throughout the project.


Antigens for successful antibody generation have to be immunogenic, as pure as possible and non-toxic for the animal. The most common immunogens are peptides or proteins, but also exosomes, whole cells expressing a surface antigen of your interest, or haptens (e.g.nucleotides or chemical substances) coupled to a carrier protein.


Proteins are the best choice for immunization as they lead to antibodies recognizing different epitopes of the protein. Ideally, the protein is purified with a cleavable tag (e.g. HA, His, Strep). Proteins without the tag can be used for immunization and tagged protein is used for screening. This way we avoid the generation of tag-specific antibodies and have the advantage of directed protein binding to the ELISA plates for screening. Proteins smaller than 10 - 15 kDa need to be conjugated to a larger carrier, e.g. GST or MBP. Approx. 1-2 mg of protein is required for immunization and screening assays.


If protein is not available, or for the generation of antibodies against post-translational modifications, peptides can be used for immunization.  Immunizing different peptides from one protein increases the chance of successful antibody generation. Peptides are selected based on their antigenicity e.g. hydrophilicity, amphipathicity, surface probability and sequence variability. We assist in choosing the best peptide(s) and can also arrange peptide ordering. Usually we use peptides that are coupled to ovalbumin for immunization and His-tagged biotinylated peptides for the subsequent screening process.

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