Carmen Sweeney, Ph.D. Method Development Department Manager Charles River Labs
The Devil’s in the Details
The purpose of a vaccine is to deliver antigen or antigens in order to induce an immune response and immune memory. Most vaccines are prophylactic or preventive and the main aim of these types of vaccines is to induce long-term immune memory which can be called upon at some stage in the future to protect the host from infection or at least to lessen the severity of disease.
How these immune responses are induced, either naturally or via a vaccine, is a complicated process however, and in fact many modern vaccines are based primarily on empirical evidence. We know they work, we just don't quite know why they work.
Closing this gap is one of the primary functions of immunogenicity tests, assays designed to measure precisely how and how well the immune system responds to a pathogen. Immunogenicity, in its simplest terms, is the ability of a particular substance, such as an antigen, to provoke an immune response. We can use the immune system to protect ourselves against new or harmful pathogens using vaccines or antitoxins. In addition to prophylactic and therapeutic benefits, the molecules of the immune system, such as antibodies, can be manipulated and used as diagnostic or biotech tools.
This immune response can be quantified and used to determine the potency (immunopotency) of vaccines, but the devil, as they say, is in the details. One major obstacle that clients face in the release of their vaccine products is the development of a robust, reliable and 'fit for purpose' immunopotency or batch release assay. In a nutshell, it is important to choose an assay that can be used on a regular and routine basis (e.g. GMP batch release). The assay also needs to perform properly and be reproducible.
Here are seven key considerations in developing an in vivo immunopotency assay:
- Remember the 3Rs. Assuming no in vitro alternative is available, have you minimized and justified the number of animals being used in the assay? What plans do you have for further animal number reduction through assay refinement?
- Choose Your Animal Model Wisely. Try to make sure the strain is readily and widely available, and find out if there are any restrictions on supply in different countries. One should also monitor the health status of the animals to ensure they are SPF, if necessary, and factor in the cost of the strain. The well-validated CD1 or BALB/c strains are stalwarts in the routine use of assays for batch release testing and are readily available. Working with less common strains and more problematic or complicated assays can require more time and money because the tests have to be done repeatedly in order to ensure that a robust, reliable and fit for purpose assay is developed that can be used on a long term basis to determine the immunopotency of the vaccine.
- Keep Your Animals Safe. To minimize contamination and exposure to nontest item antigens, be sure to house and handle animals in facilities that have adequate biosecurity measures in place, e.g., IVC cage-changing stations, biological safety cabinets/work stations. Staff should wear appropriate personal protective equipment. Buildings that conduct immunogenicity-based assays should be designed to ensure minimal animal exposure to contamination, be able to contain the bioiological safety level of the test item and offer appropriate personnel controls.
- Handle Reference Materials Carefully. Is the vaccine product classified as a pathogen or a genetically modified microorganism (GMM) and are there special licensing requirements for handling, testing, and shipment of the test or reference material? For example, import licences are required for international shipments of pathogenic and GMM material. In addition, approval and licensing for the handling and testing of GMM material is usually required, e.g., EPA license needs to be sought.
- Be Attentive to the Regimen…and Who Administers It. Before performing an immunopotency test one should always account for the dosing level of the vaccine and whether an adjuvant is necessary to ensure an adequate antibody response. Also relevant during the analysis is the number of doses required, e.g,. is a single administration sufficient or is one or more boosters required. The timing of these administrations is also critical to determine. It goes without saying that having adequately-trained staff performing the tests is essential to ensuring consistent delivery of the test item.
- Choose the Right Assay. If you don't have a reasonably good idea which endpoint analysis, such as ELISA, serum neutralization, plaque reduction, offers the most reproducible assay during batch release testing, your hard work will falter. One way to avoid this is by knowing the true mode of action of your vaccine product. It is usually necessary to develop a minimum of two endpoint assays and then selection of the candidate endpoint assay for batch release will be based on data and will ensure that the assay is reproducible and reliable.
- Don't Overtax Your Lab. If it becomes unwieldy handling the volume of routine batch release testing required in-house, you might need to outsource the work. Whether to do this before or after validation/qualification of the assay will come down to whether you have the experience and knowledge available in house to design and execute the qualification/validation study or if it would be more appropriate to out-source this final aspect of assay development to a CRO.
Carmen Sweeney, Ph.D. (firstname.lastname@example.org), is the method development department manager for Charles River's preclinical services site in Ballina, Ireland. This article previously appeared on Charles River's science blog Eureka.