Vaccine 35:1197C1199. Commons Attribution 4.0 International license. FIG?S4. Timeline for survival study of cPVR mice. Download FIG?S4, PDF file, 0.1 MB. Copyright ? 2018 Radicicol Shin et al. This content is distributed under the terms of the Creative Commons Attribution 4.0 International license. FIG?S5. Determination of optimal time for testing neutralizing antibody titer. Download FIG?S5, PDF file, 0.02 MB. Copyright Mouse monoclonal to INHA ? 2018 Shin et al. This content is distributed under the terms of the Creative Commons Attribution 4.0 International license. ABSTRACT As oral poliovirus vaccine (OPV) causes vaccine-associated paralytic poliomyelitis, the polio endgame strategy introduced by the Global Polio Eradication Initiative calls for a phased withdrawal of OPV and an introduction of inactivated poliovirus vaccine (IPV). The introduction of IPV creates challenges in maintaining the cold chain for vaccine storage and distribution. Recent advances in lyophilization have helped in finding a temperature-stable formulation for multiple vaccines; however, poliovirus vaccines have yet to capture a stable, safe formula for lyophilization. In addition, efficient methods for antigen measurement are needed for screening stable vaccine formulations. Here, we report size exclusion high-performance liquid chromatography (SE-HPLC) as a reliable means to identify the leading lyophilized formulation to generate thermostable Sabin inactivated poliovirus vaccine (sIPV). High-throughput screening and SE-HPLC determined the leading formulation, resulting in 95% D-antigen recovery and low residual moisture content of sIPV following lyophilization. Furthermore, the lyophilized sIPV remained stable after 4 weeks of incubation at ambient temperature and induced strong neutralizing antibodies and full protection of poliovirus receptor transgenic mice against the challenge of wild-type poliovirus. Overall, this report describes a novel means for the high-throughput evaluation of sIPV antigenicity and a thermostable lyophilized sIPV with vaccine potency. method to evaluate vaccine potency. The potency of IPV determined by the assay is expressed in arbitrarily defined D-antigen units (D-AgU). The D-AgU was established in the early 1960s (22) following characterization of purified virus preparations by sucrose gradient centrifugation where two bands were identified. One, the D fraction (D-antigen), was associated with infectious virus with intact structure as revealed by electron microscopy and RNA content. The other, the C fraction (C-antigen), contained low infectivity with little RNA and possessed a structure that was similar to the structure of the heat-treated virus. As induction of neutralizing antibodies is associated with the immunization of intact virus structures (D-antigen) but not with the immunization of C-antigen viral preparations, the potency of IPV has been a function of the D-antigen content. Thus, efficient methods for D-antigen measurement are needed for screening stable vaccine formulations. In this study, Radicicol various surfactant-based formulations were screened for Sabin inactivated poliovirus vaccine (sIPV) Radicicol lyophilization, and size exclusion high-performance liquid chromatography (SE-HPLC) (23) was implemented as a novel high-throughput formulation assay for D-antigen quantitation of sIPV. Finally, a room-temperature-stable sIPV prepared by leading formulation induced strong neutralizing antibodies and full protection against wild-type (WT) poliovirus challenge (%)(%)= 0)protective efficacy testing against wild-type (WT) PV infection (31). Open in a separate window FIG 3 Lyophilized sIPV remains stable at elevated temperatures. To test the thermostability of lyophilized sIPV, lyophilized sIPV from formulations F4, F8, and F9 was incubated at different temperatures and D-Ag recovery was measured using conventional ELISA. (A) D-Ag unit recovery over 4?weeks of incubation at 4C. (B) D-Ag unit recovery over 4?weeks of incubation at 25C. (C) D-Ag unit recovery over 4?weeks of incubation in 40C. A total of six groups of poliovirus receptor transgenic (cPVR) mice (= 8) expressing human CD155 for viral entry (32, 33) were vaccinated with 20 D-AgU of sIPV, lyophilized (lyo) sIPV, IPOL-IPV (a trivalent polio vaccine distributed by Sanofi Pasteur), or phosphate-buffered saline (PBS) incubated for 4?weeks at the indicated temperatures. cPVR mice were vaccinated, boosted, challenged, and observed for 2 weeks for signs of paralysis (Fig.?S4), using a blind scoring method outlined in the WHO standard operating procedure for OPV neurovirulence testing (34). For the serum neutralizing titers, we first checked day ?1 serum and confirmed that no mice had seroconverted, as the neutralization titers were below the detection limit (Fig.?S5). We then checked the neutralization titers on days 13 and 21 and found that the neutralization titers measured on day 13 were approximately 2 logs lower than those measured on day 21 (Fig.?S5). On the basis of the standardized WHO potency testing of IPV in rodents along with the results of the experiment whose results are shown in Fig.?S5, day 21 after vaccination was determined as a time point for the neutralization.
