Evaluation of the rapid analyte measurement platform (RAMP) for the detection of Bacillus anthracis at a crime scene

Hoile, Rebecca Jane; Yuen, Marion; James, Gregory; Gilbert, Gwendolyn L.

doi:10.1016/j.forsciint.2006.09.004

Cited by 15 publications

(8 citation statements)

References 6 publications

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“…For example, immunoassays targeting either spore antigens or the anthrax toxins can be completed within 15–120 min, which is comparable with the signal response time described in this report (Mabry et al. 2006; Hoile et al. 2007).…”

Section: Discussionsupporting

confidence: 73%

Phage-mediated bioluminescent detection ofBacillus anthracis

Schofield

Westwater

2009

Journal of Applied Microbiology

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Aims: Bacillus anthracis, the causative agent of anthrax, is a serious human pathogen. The aim of this study was to provide the proof of principle results for the development of a ‘bioluminescent’ reporter bacteriophage that was capable of specifically detecting B. anthracis. Methods and Results: The reporter phage was engineered by integrating the bacterial luxA and luxB reporter genes into a nonessential region of the lysogenic Wβ phage genome. This resulted in a phage that was capable of specifically infecting and conferring a bioluminescent phenotype to B. anthracis viable cells. No processing or cell preparation was required; the phage and cells were simply mixed, and the samples were analysed for bioluminescence. A bioluminescent signal was evident after 16 min postinfection of vegetative cells. The strength and time required to generate the signal was dependent on the number of cells present. Nevertheless, 103 CFU ml−1 was detectable within 60 min. The utility of the bioluminescent phage was analysed using spores as the starting material. The Wβ::luxAB phage was able to transduce a bioluminescent signal to germinating spores within 60 min. Conclusions: This proof of principle study demonstrates that the reporter phage displays promise as a tool for the rapid detection of B. anthracis. Significance and Impact of the Study: The new methodology offers the potential for the detection of viable cells from either environmental or clinical samples.

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Section: Discussionsupporting

confidence: 73%

Phage-mediated bioluminescent detection ofBacillus anthracis

Schofield

Westwater

2009

Journal of Applied Microbiology

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“…The sensitivity of spore detection by the Luminex assay was substantially increased compared to the corresponding capture ELISA, with the detection of 10 3 to 10 4 spores per ml. Currently available immunological detection systems offer higher detection limits (11,12,14). The platform can be also used in multiplex assays, and we are currently evaluating the simultaneous detection of different biothreat bacteria in mixed samples.…”

Section: Discussionmentioning

confidence: 99%

Anthrax Spore Detection by a Luminex Assay Based on Monoclonal Antibodies That Recognize Anthrose-Containing Oligosaccharides

Tamborrini

Holzer

Seeberger

et al. 2010

Clin Vaccine Immunol

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The similarity of endospore surface antigens between bacteria of the Bacillus cereus group complicates the development of selective antibody-based anthrax detection systems. The surface of B. anthracis endospores exposes a tetrasaccharide containing the monosaccharide anthrose. Anti-tetrasaccharide monoclonal antibodies (MAbs) and anti-anthrose-rhamnose disaccharide MAbs were produced and tested for their fine specificities in a direct spore enzyme-linked immunosorbent assay (ELISA) with inactivated spores of a broad spectrum of B. anthracis strains and related species of the Bacillus genus. Although the two sets of MAbs had different fine specificities, all of them recognized the tested B. anthracis strains and showed only a limited cross-reactivity with two B. cereus strains. The MAbs were further tested for their ability to be implemented in a highly sensitive and specific bead-based Luminex assay. This assay detected spores from different B. anthracis strains and two cross-reactive B. cereus strains, correlating with the results obtained in direct spore ELISA. The Luminex assay (detection limit 10 3 to 10 4 spores per ml) was much more sensitive than the corresponding sandwich ELISA. Although not strictly specific for B. anthracis spores, the developed Luminex assay represents a useful first-line screening tool for the detection of B. anthracis spores.

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“…Such assays have not been well described before. Besides PCR-based detection, only a few immunological detection systems, such as lateral-flow immunoassays, are available, but they offer detection limits that range from only 10 5 to 10 6 spores/ml (12,19). Unfortunately, no information on the target antigens of the antibodies used in those assays is available.…”

Section: Discussionmentioning

confidence: 99%

“…When the mail system was used for biological attacks in the United States in 2001, the great efficacy of this pathogen to infect many people when it was intentionally distributed became visible (16). In addition to PCR techniques, which could be hampered by inhibitory factors, and inefficient means for the preparation of DNA from spores, the rapid immunological detection of B. anthracis from complex specimens is currently possible only when the spore concentrations are relatively high and when handheld test kits are used (12). In cases of lower concentrations, time-consuming cultivation and later identification would take too long in an emergency situation (11).…”

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confidence: 99%

Development of Antibodies against Anthrose Tetrasaccharide for Specific Detection of Bacillus anthracis Spores

Kuehn

Kòváč

Saksena

et al. 2009

Clin Vaccine Immunol

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Methods for the immunological detection ofThe BclA protein is a major component of the exosporium of B. anthracis spores and is decorated by the tetrasaccharide indicated above. The anthrose-containing tetrasaccharide chain seems to be highly specific for B. anthracis, which makes it a key biomarker for the detection of these spores. The different immunizations led to anthrose-reactive polyclonal and monoclonal antibodies which were analyzed by various methods to characterize their ability to discriminate between B. anthracis and other Bacillus spp. Multiple applications, such as enzyme-linked immunosorbent assay, indirect immunofluorescence assay, and electron microscopy, revealed the specificities of the polyclonal and monoclonal antibodies generated for B. anthracis spore detection. All polyclonal antibodies were able to correctly identify the B. anthracis strains tested and showed only minimal cross-reactivities with other Bacillus strains. Moreover, the antibodies generated proved functional in a new capture assay for B. anthracis spores and could therefore be useful for the detection of spores in complex samples.

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Evaluation of the rapid analyte measurement platform (RAMP) for the detection of Bacillus anthracis at a crime scene

Cited by 15 publications

References 6 publications

Phage-mediated bioluminescent detection ofBacillus anthracis

Phage-mediated bioluminescent detection ofBacillus anthracis

Anthrax Spore Detection by a Luminex Assay Based on Monoclonal Antibodies That Recognize Anthrose-Containing Oligosaccharides

Development of Antibodies against Anthrose Tetrasaccharide for Specific Detection of Bacillus anthracis Spores

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