Single-dilution assay of new vaccines produced using plant and insect cell substrates

Large numbers of animals are required in multi-dilution assays of vaccines produced in mammalian, plant and insect cell substrates. Animal vaccination and serum sampling require skilled labour, adding to testing costs. More tests are required with homologous reference preparations. Use of a single antigen reference to assay combined formulations is invalid due to synergistic or other effects of multiple antigens. Licensing authorities in Europe recommend use of single-dilution assays for some vaccines, together with homologous reference preparations. Vaccine and reference formulations in a single-dilution assay must have similar compositions to ensure similar dose response relationships. Design and validation of such a single-dilution assay is the main consideration of this article. Multi-dilution in vivo vaccine assays have inherent problems also affecting single-dilution assays. Statistical analysis of data from multi-dilution assays requires significance tests of assay validity, but with a single-dilution assay, some aspects of validity are assumed. These aspects include linearity and parallelism of the dilution-response curves for both reference and test preparations. This article shows that despite inherent problems associated with multi-dilution animal assays, the need to move to a simpler single-dilution model necessitates establishing minimal inherent conditions in the multi-dilution assay for a valid interpretation of the single-dilution assay.