Comparability of ELISA and toxin neutralization to measure immunogenicity of Protective Antigen in mice, as part of a potency test for anthrax vaccines.
ABSTRACT Complexities of lethal challenge models have prompted the investigation of immunogenicity assays as potency tests of anthrax vaccines. An ELISA and a lethal toxin neutralization assay (TNA) were used to measure antibody response to Protective Antigen (PA) in mice immunized once with either a commercial or a recombinant PA (rPA) vaccine formulated in-house. Even though ELISA and TNA results showed correlation, ELISA results may not be able to accurately predict TNA results in this single immunization model.
- SourceAvailable from: Okechukwu Ndumnego[Show abstract] [Hide abstract]
ABSTRACT: Presently, few data exist on the level and duration of anti-protective antigen (PA) IgG in vaccinated livestock. Various adaptation of enzyme-linked immunosorbent assays (ELISAs) have been developed in studies to assess immune response following vaccination, albeit mostly in laboratory rodent models. The quantitative anti-anthrax IgG ELISA in this study describes a method of enumerating the concentration of anti-PA specific IgG present in sera of immunized goats, with the aid of an affinity-purified caprine polyclonal anti-anthrax PA-83 IgG standard. This was compared with the anthrax toxin neutralization assay (TNA) which measures a functional subset of toxin neutralizing anti-PA IgG. The measured concentrations obtained in the standard curve correlated with the known concentration at each dilution. Percentage recovery of the standard concentrations ranged from 89 to 98% (lower and upper asymptote respectively). Mean correlation coefficient (r2) of the standard curve was 0.998. Evaluation of the intra-assay coefficient of variation showed ranges of 0.23-16.90% and 0.40-12.46% for days 28 and 140 sera samples respectively, following vaccination. The mean inter-assay coefficient of variation for triplicate samples repeated on 5 different days was 18.53 and 12.17% for days 28 and 140 sera samples respectively. Spearman's rank correlation of log-transformed IgG concentrations and TNA titres showed strong positive correlation (rs = 0.942; p = 0.01). This study provides evidence that an indirect ELISA can be used for the quantification of anti-anthrax PA IgG in goats with the added advantage of using single dilutions to save time and resources. The use of such related immunoassays can serve as potential adjuncts to potency tests for Sterne and other vaccine types under development in ruminant species. This is the first report on the correlation of polyclonal anti-anthrax PA83 antibody with the TNA in goats.BMC Veterinary Research 12/2013; 9(1):265. DOI:10.1186/1746-6148-9-265 · 1.74 Impact Factor
- [Show abstract] [Hide abstract]
ABSTRACT: We have developed a rapid, reliable, and sensitive quantitative flow cytometric assay to measure the in vitro potency and stability of DNA vaccines to be delivered either by particle-mediated epidermal delivery (PMED) or by electroporation. The method involves transfecting cells with test DNA and comparing the measured antigen expression to that generated with expression from known quantities of reference material DNA. The assay was adapted for performance under Good Laboratory Practice (GLP) guidelines and was successfully utilized to perform potency testing in support of a Phase I study for two hantavirus DNA vaccines delivered by gene gun. The results from the potency assays conducted over a 24-month period using this method proved to be highly reproducible with high signal-to-noise ratios. The assay was also adapted to assess the in vitro potency and stability of a DNA vaccine for Venezuelan equine encephalitis virus that will be delivered by electroporation. Our results indicate that this assay can be readily applied to support potency and stability testing of numerous DNA vaccines delivered by various methods, including multiagent vaccines.Vaccine 09/2011; 29(39-29):6728-6735. DOI:10.1016/j.vaccine.2010.12.053 · 3.49 Impact Factor
- Oral Oncology 07/2011; 47. DOI:10.1016/j.oraloncology.2011.06.087 · 3.03 Impact Factor