Antigen / Immunogen Discovery and Optimization

Step 1: Screen for binding to virus-specific antibody

Phage display is a powerful in vitro method that can be used to screen millions of antibodies against viral antigens, which are then considered as potential vaccine candidates.
 

Step 2: Immunogen characterization

Immunogenicity studies are conducted to determine whether the antigens identified in the previous step will initiate an immune response, since phage display only screens for antigen-antibody binding. If the antigen evokes an immune response (often measured by an ELISA), the antigen, now considered an immunogen, will move forward. Additional immunogen characterization is used to engineer desirable properties into the immunogen in the next stage.
 

Step 3: Protein (immunogen) engineering

Some properties of immunogens are known to cause undesirable effects when injected into animals or humans. Therefore, genetic engineering tools like CRISPR/Cas9 are used to manipulate the DNA encoding the immunogen so that they no longer possess those properties.
 

Step 4: Cell transfection and cloning

Once the DNA has been optimized, it is necessary to scale up the production of the immunogen for further testing. At this stage, it is critical to generate a high expressing, clonally derived cell line. This process often involves genetic engineering techniques described above along with single cell cloning into microwell plates, particularly if using mammalian cell lines.
 

Step 5: Screen for quality attributes including titer

Following single cell cloning, cell lines are monitored for growth and assessed for protein expression. The immunogen is then purified and prepared for injection into an animal in order to induce an immune response. Antibodies generated from the animal are then screened in a virus neutralization assay to determine vaccine efficacy.