Qualification and performance characteristics of a quantitative enzyme-linked immunosorbent assay for human lgG antibodies to anthrax lethal factor antigen

Vi polysaccharide from Salmonella enterica serotype Typhi is used as one of the available vaccines to prevent typhoid fever. Measurement of Vi-specific serum antibodies after vaccination with Vi polysaccharide by enzyme-linked immunosorbent assay (ELISA) may be complicated due to poor binding of the Vi polysaccharide to ELISA plates resulting in poor reproducibility of measured antibody responses. We chemically conjugated Vi polysaccharide to fluorescent beads and performed studies to determine if a bead-based immunoassay provided a reproducible method to measure vaccine-induced anti-Vi serum IgG antibodies. Compared to ELISA, the Vi bead immunoassay had a lower background and therefore a greater signal-to-noise ratio. The Vi bead immunoassay was used to evaluate serum anti-Vi IgG in 996 subjects from the city of Kolkata, India, before and after vaccination. Due to the location being one where Salmonella serotype Typhi is endemic, approximately 45% of the subjects had protective levels of anti-Vi serum IgG (i.e., 1 g/ml anti-Vi IgG) before vaccination, and nearly 98% of the subjects had protective levels of anti-Vi serum IgG after vaccination. Our results demonstrate that a bead-based immunoassay provides an effective, reproducible method to measure serum anti-Vi IgG responses before and after vaccination with the Vi polysaccharide vaccine.

Section: Discussionsupporting

confidence: 90%

Development of a Bead Immunoassay To Measure Vi Polysaccharide-Specific Serum IgG after Vaccination with the Salmonella enterica Serovar Typhi Vi Polysaccharide

Staats

Kirwan

Whisnant

et al. 2010

“…Two immunoassays are currently in routine use for the quantitation of antibodies against PA: an enzyme-linked immunosorbent assay (ELISA) that measures antibody to PA (21,24) and an LT toxin neutralization assay (TNA) (3,6,17). The ELISA was developed in parallel for use with multiple species; however, the species specificity of the ELISA limits direct comparison of the responses between different animal species as well as direct comparison of the responses between animals and humans.…”

mentioning

confidence: 99%

Interlaboratory Comparison of Results of an Anthrax Lethal Toxin Neutralization Assay for Assessment of Functional Antibodies in Multiple Species

Omland

Brys

Lansky

et al. 2008

The anthrax lethal toxin neutralization assay (TNA) will likely be used to correlate the protection offered by new anthrax vaccines in animal models to the immunogenicity that will be provided in humans. TNA data are being generated in several different laboratories to measure the immune responses in rabbits, nonhuman primates, and humans. In order to compare data among species and laboratories, a collaborative study was conducted in which 108 samples from the three species were analyzed in seven independent laboratories. Six of the seven laboratories had participated in an interlaboratory technology transfer of the TNA. Analysis of the titration curves generated by samples from each species indicated that the behaviors of the samples from all species were similar; the upper and lower asymptotes and the slopes of the curves were less than 30% divergent from those for human reference material. Dilutional linearity was consistent among samples from each species, with spike to effective dilution at 50% inhibition (ED 50 ) slopes of less than 1.2 for all species. Agreement among the laboratories with consensus values was within 10% of the ED 50 s for all samples and within 7.5% of the quotients of the test sample ED 50 and the reference standard ED 50 (NF 50 s) for all samples. The relative standard deviations obtained when data from all laboratories and for all species were combined were 45% for the ED 50 s and 35% for the NF 50 s. These precision data suggest that the NF 50 readout may normalize the values generated by different laboratories. This study demonstrates that the TNA is a panspecies assay that can be performed in several different laboratories with a high degree of quantitative agreement and precision.

“…As well, the number of priming doses may be reduced (J. (26). With this, they have elucidated the antibody response and anticipate an assessment of the contribution of the LF-specific immune response to protection against anthrax.…”

Section: Discussionmentioning

confidence: 99%

Rapid Point-of-Care Test To Detect Broad Ranges of Protective Antigen-Specific Immunoglobulin G Concentrations in Recipients of the U.S.-Licensed Anthrax Vaccine

Bienek

Biagini

Charlton

et al. 2008

, Clin. Vaccine Immunol. 13:541-546, 2006), we investigated the agreement between a validated anthrax protective antigen (PA) immunoglobulin G (IgG) enzyme-linked immunosorbent assay (ELISA) and the LFA for 335 unvaccinated and vaccinated subjects. We also investigated the performance of the LFA under the following conditions: thermal shock (i.e., thermal cycling between temperature extremes), high temperature/high relative humidity, high temperature/low relative humidity, and low temperature/low relative humidity. With the anti-PA ELISA used as a standard, the LFA was shown to be optimally diagnostic at 11 g/ml anti-PA-specific IgG. At this concentration, the LFA specificity and sensitivity were 98% (95% confidence interval [CI], 97% to 100%) and 92% (CI, 88% to 97%), respectively. Receiver operating characteristic curve analysis yielded an area under the curve value of 0.988 (CI, 0.976 to 1.00), suggesting that the LFA is an extremely accurate diagnostic test. For <4 or >50 g/ml PA-specific IgG, the LFA results for each environmental condition were identical to those obtained in the laboratory. These data indicate that this rapid point-of-care test would be a feasible tool in monitoring the serological antibody responses of individuals that have been vaccinated against anthrax.