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Three Competitive ELISAs to Quantify the D-Antigen Content of Aluminum-Salt Adjuvanted Recombinant Polio VLPs (Types
Yanli Liu1, John M Hickey1, Geetha Satya Sainaga Jyothi Vaskuri1
1Department of Pharmaceutical Chemistry, Vaccine Analytics and Formulation Center, University of Kansas, 2030 Becker Drive, Lawrence, KS 66047, USA.
Background/Objectives:
Recombinant poliovirus (PV) virus-like particle (VLP) antigens mimic the conformation of the surface proteins in native PVs (i.e., serotype-specific D-antigen epitopes). Since they lack genomes and are non-infectious, PV-VLPs offer the promise of a safer, next-generation polio vaccine compared to traditional inactivated (IPV) or attenuated live (OPV) vaccines. Sandwich D-antigen ELISA formats are commonly used to measure the in vitro potency values (relative D-antigen content, DU/mL) of unadjuvanted trivalent IPV antigens. If IPV is formulated with aluminum-salt adjuvants, however, a pretreatment step (i.e., adjuvant dissolution or antigen desorption) is required, which may compromise antigen integrity during sample handling.
Methods:
This work describes the development of three competitive ELISAs to measure the relative D-antigen content of aluminum-salt adjuvanted PV-VLPs (Types 1, 2, 3) without the need for pretreatment.
Results:
First, key assay parameters were established, including specificity, accuracy, precision, linearity, limit of quantification, and stability-indication. Next, preformulation characterization studies were performed with these methods including (1) rank-ordering the inherent thermal stability profiles of the PV-VLPs (Types 1 > 3 > 2) in-solution and adsorbed to an aluminum phosphate adjuvant (AdjuPhos™, AP) and (2) determining the effect of formulation variables on the thermal stability profiles of AP-adsorbed PV-VLPs including antimicrobial preservatives (thimerosal, 2-PE) and five different antigens present in pediatric combination vaccines (D, T, wP, Hib, Hep B).
Conclusions:
The development and application of three competitive D-antigen ELISAs were demonstrated, and future use in formulation and storage stability studies with the AP-adjuvanted, trivalent PV-VLPs (Types 1, 2, 3) is discussed with the long-term goal to develop a stable, efficacious, multi-dose, hexavalent combination vaccine presentation.
