Validation of the anthrax lethal toxin neutralization assay

Hering, Donna; Thompson, William L.; Hewetson, John F.; Little, Stephen F.; Norris, Sarah L.; Pace-Templeton, Judith G

doi:10.1016/j.biologicals.2003.09.003

Cited by 82 publications

(74 citation statements)

References 5 publications

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“…Both have been well described in the literature and have been validated. 16,17 Briefly, the ELISA quantifies the level of anti-PA-IgG and reports concentrations in mg/mL. , is added to wells and cell viability is determined colorimetrically.…”

Section: Methodsmentioning

confidence: 99%

Safety, Reactogenicity, and Immunogenicity of a Recombinant Protective Antigen Anthrax Vaccine Given to Healthy Adults

Campbell¹,

Clement²,

Wasserman³

et al. 2007

Human Vaccines

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Background: Bacillus anthracis causes anthrax, a vaccine-preventable zoonotic disease that may follow intentional or unintentional exposure to its spores. Although an anthrax vaccine is currently licensed in the USA, better vaccines are desirable for both pre-and post-exposure prophylaxis.Methods: Healthy adults, aged 18-40 years, received anthrax immunization with either licensed Anthrax Vaccine Adsorbed (AVA, BioThrax™), or an experimental recombinant Protective Antigen vaccine (rPA) produced from an avirulent, non-spore-forming strain of B. anthracis at one of four doses (5, 25, 50 or 75 mg). Volunteers were followed for safety, reactogenicity, and immunogenicity.Results: rPA vaccine was well tolerated with a low rate of local or systemic reactions. Although antibody responses were poor following unadjuvanted rPA administration, 89 and 100% of volunteers who received Alhydrogel-adjuvanted rPA given intramuscularly had four-fold increases by enzyme-linked immunosorbent and toxin neutralization assays, respectively. Peak antibody responses to adjuvanted rPA given intramuscularly were equivalent to AVA, given either intramuscularly or subcutaneously, when measured by either assay.Conclusions: This recombinant Protective Antigen anthrax vaccine, when given with the adjuvant Alhydrogel to healthy adults in two intramuscular injections four weeks apart, is very well-tolerated and highly immunogenic.

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Section: Methodsmentioning

confidence: 99%

Safety, Reactogenicity, and Immunogenicity of a Recombinant Protective Antigen Anthrax Vaccine Given to Healthy Adults

Campbell¹,

Clement²,

Wasserman³

et al. 2007

Human Vaccines

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show abstract

“…Samples were retrieved from −70 • C storage that met preestablished criteria: (a) Vaccinations must have occurred within defined time intervals after receipt of the initial AVA injection (day 0 [dose #1], day 14 [range [11][12][13][14][15][16][17][18][19][20][21] ). (b) Pre-injection sera were defined as those taken on the day of each vaccine dose.…”

Section: Methodsmentioning

confidence: 99%

“…anthracis lethal toxin neutralization activity was measured by using a colorimetric toxin neutralization assay (TNA) [10,11]. In brief, confluent monolayers of J774A.1 (mouse macrophage) cells were grown in 96-well plates and used after overnight incubation.…”

Section: Laboratory Studiesmentioning

confidence: 99%

Patterns of antibody response in humans to the anthrax vaccine adsorbed (AVA) primary (six-dose) series☆

Pittman

Norris

Oro

et al. 2006

Vaccine

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“…Lethal toxin neutralization titers were measured as the ability of the sera to block the cytotoxicity of lethal toxin in a J774A.1 macrophage cell line (passage 3-20) as described by Hering et al [32] with some modifications as described below. Antisera diluted in cell culture medium were added in duplicate to 96-well tissue culture-treated flat microtiter plates (Corning costar, Acton, MA) that were pre-incubated with purified recombinant PA and LF at a final concentration of 50 ng/ml each, for 1 h at 37°C.…”

Section: Immunogenicity Studiesmentioning

confidence: 99%

Multicomponent anthrax toxin display and delivery using bacteriophage T4

Shivachandra

Peachman

et al. 2007

Vaccine

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We describe a multicomponent antigen display and delivery system using bacteriophage T4. Two dispensable outer capsid proteins, Hoc (highly antigenic outer capsid protein, 155 copies) and Soc (small outer capsid protein, 810 copies), decorate phage T4 capsid. These proteins bind to the symmetrically localized capsid sites, which appear following prohead assembly and expansion. We hypothesized that multiple antigens fused to Hoc can be displayed on the same capsid and such particles can elicit broad immunological responses. Anthrax toxin proteins, protective antigen (PA), lethal factor (LF), and edema factor (EF), and their functional domains, were fused to Hoc with an N-terminal hexa-histidine tag and the recombinant proteins were over-expressed in E. coli and purified. Using a defined in vitro assembly system, the anthrax-Hoc fusion proteins were efficiently displayed on T4 capsid, either individually or in combinations. All of the 155 Hoc binding sites can be occupied by one antigen, or they can be split among two or more antigens by varying their molar ratio in the binding reaction. Immunization of mice with T4 phage carrying PA, LF, and EF elicited strong antigen-specific antibodies against all antigens as well as lethal toxin neutralization titers. The triple antigen T4 phage elicited stronger PA-specific immune responses than the phage displaying PA alone. These features offer novel avenues to develop customized multicomponent vaccines against anthrax and other pathogenic diseases.

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Validation of the anthrax lethal toxin neutralization assay

Cited by 82 publications

References 5 publications

Safety, Reactogenicity, and Immunogenicity of a Recombinant Protective Antigen Anthrax Vaccine Given to Healthy Adults

Safety, Reactogenicity, and Immunogenicity of a Recombinant Protective Antigen Anthrax Vaccine Given to Healthy Adults

Patterns of antibody response in humans to the anthrax vaccine adsorbed (AVA) primary (six-dose) series☆

Multicomponent anthrax toxin display and delivery using bacteriophage T4

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