Monoclonal antibodies against spore antigens of Bacillus anthracis

Phillips, A

doi:10.1016/0378-1097(88)90028-6

Cited by 8 publications

(11 citation statements)

References 21 publications

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“…A possible explanation for this effect is damage to B. anthracis spores during gamma irradiation, allowing increased access to target chromosomal DNA. This explanation is consistent with a study conducted by Phillips et al (31), which reported that gamma irradiation resulted in structural damage of B. anthracis spores. In contrast, irradiation did not significantly affect detection of the pXO1 and pXO2 plasmids of B. anthracis by real-time PCR.…”

Section: Discussionsupporting

confidence: 82%

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Gamma Irradiation Can Be Used To Inactivate Bacillus anthracis Spores without Compromising the Sensitivity of Diagnostic Assays

Dauphin

Newton

Rasmussen

et al. 2008

Appl Environ Microbiol

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The use of Bacillus anthracis as a biological weapon in 2001 heightened awareness of the need for validated methods for the inactivation of B. anthracis spores. This study determined the gamma irradiation dose for inactivating virulent B. anthracis spores in suspension and its effects on real-time PCR and antigen detection assays. Strains representing eight genetic groups of B. anthracis were exposed to gamma radiation, and it was found that subjecting spores at a concentration of 10 7 CFU/ml to a dose of 2.5 ؋ 10 6 rads resulted in a 6-log-unit reduction of spore viability. TaqMan real-time PCR analysis of untreated versus irradiated Ames strain (K1694) spores showed that treatment significantly enhanced the detection of B. anthracis chromosomal DNA targets but had no significant effect on the ability to detect targets on the pXO1 and pXO2 plasmids of B. anthracis. When analyzed by an enzyme-linked immunosorbent assay (ELISA), irradiation affected the detection of B. anthracis spores in a direct ELISA but had no effect on the limit of detection in a sandwich ELISA. The results of this study showed that gamma irradiation-inactivated spores can be tested by real-time PCR or sandwich ELISA without decreasing the sensitivity of either type of assay. Furthermore, the results suggest that clinical and public health laboratories which test specimens for B. anthracis could potentially incorporate gamma irradiation into sample processing protocols without compromising the sensitivity of the B. anthracis assays.

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Section: Discussionsupporting

confidence: 82%

“…It has been reported that gamma irradiation of B. anthracis spores may affect epitopes recognized by MAbs and PAbs (7,31). The results of this study are consistent with this observation, as the detection level of irradiated spores was decreased when the MAb was used and enhanced when the PAb was used.…”

Section: Discussionsupporting

confidence: 81%

Gamma Irradiation Can Be Used To Inactivate Bacillus anthracis Spores without Compromising the Sensitivity of Diagnostic Assays

Dauphin

Newton

Rasmussen

et al. 2008

Appl Environ Microbiol

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“…4). Detection and identification of Bacillus anthracis spores using flow cytometry has been demonstrated earlier using formalin-fixed spores 5,7,13 . Stopa 14 has reported the use of live as well as gamma-irradiated spores for detection and identification of Bacillus anthracis spores.…”

Section: Resultsmentioning

confidence: 97%

“…Moreover, culture methods take 18-24 h and characterisation is done through a series of biochemical tests 4 . The development of polymerase chain reaction (PCR) and antibody-based systems like immunoradiometric assay (IRMA), enzyme-linked immunosorbent assay (ELISA), immuno-fluorescence, and immunochemiluminescence for the detection of Bacillus anthracis have been investigated by several workers [5][6][7][8][9][10] . Most of these methods are very sensitive but the two major drawbacks are the slow speed of detection, and cross-reactivity with other related Bacillus species.…”

Section: Introductionmentioning

confidence: 99%

Flow-cytometric Analysis of Bacillus anthracis Spores

Kamboj

Agarwal

Dwarakanath

et al. 2006

DSJ

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Flow-cytometric technique has been established as a powerful tool for detection and identification of microbiological agents. Unambiguous and rapid detection of Bacillus anthracis spores has been reported using immunoglobulin G-fluorescein isothiocyanate conjugate against live spores. In addition to the high sensitivity, the present technique could differentiate between spores of closely related species, eg, Bacillus cereus and Bacillus subtilis using fluorescence intensity. The technique can be used for detection of live as well as inactivated spores making it more congenial for screening of suspected samples of bioterrorism.

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“…Using two different IgG molecules against different determinants of the spore might result in more significant improvement of the selectivity. This can be achieved by further developing monoclonal antibodies against different spore epitopes of B. anthracis (23). Such reagents, labeled with a donor-acceptor pair, may produce a even more isolated gate for the target spores.…”

Section: Discussionmentioning

confidence: 99%

Detection of Frequency Resonance Energy Transfer Pair on Double-Labeled Microsphere and Bacillus anthracis Spores by Flow Cytometry

Zahavy¹,

Fisher

Bromberg³

et al. 2003

Appl Environ Microbiol

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Development of an ultrasensitive biosensor for biological hazards in the environment is a major need for pollutant control and for the detection of biological warfare. Fluorescence methods combined with immunodiagnostic methods are the most common. To minimize background noise, arising from the unspecific adsorption effect, we have adapted the FRET (frequency resonance energy transfer) effect to the immunofluorescence method. FRET will increase the selectivity of the diagnosis process by introducing a requirement for two different reporter molecules that have to label the antigen surface at a distance that will enable FRET. Utilizing the multiparameter capability of flow cytometry analysis to analyze the double-labeling/FRET immunostaining will lead to a highly selective and sensitive diagnostic method. This work examined the FRET interaction of fluorescence-labeled avidin molecules on biotin-coated microspheres as a model system. As target system, we have used labeled polyclonal antibodies on Bacillus anthracis spores. The antibodies used were purified immunoglobulin G (IgG) molecules raised in rabbits against B. anthracis exosoporium components. The antibodies were fluorescence labeled by a donor-acceptor chromophore pair, alexa488 as a donor and alexa594 as an acceptor. On labeling the spores with alexa488-IgG as a donor and alexa594-IgG as an acceptor, excitation at 488 nm results in quenching of the alexa-488 fluorescence (E q ‫؍‬ 35%) and appearance of the alexa594 fluorescence (E s ‫؍‬ 22%), as detected by flow cytometry analysis. The FRET effect leads to a further isolated gate (FL1/FL3) for the target spores compared to competitive spores such as B. thuringiensis subsp. israelensis and B. subtilis. This new approach, combining FRET labeling and flow cytometry analysis, improved the selectivity of the B. anthracis spores by a factor of 10 with respect to B. thuringiensis subsp. israelensis and a factor of 100 with respect to B. subtilis as control spores.Development of an ultrasensitive biosensor for biological hazardous in the environment is a major need for pollutant control and for the detection of biological warfare (16,22). The pathogen bacteria Bacillus anthracis is one of the biological agents likely to be used as a bioweapon (7); hence, it is the focus of a major effort in the field of rapid detection and identification (15). Fluorescence methods combined with immunodiagnostic methods are widely used for the detection and identification for biological hazards in the laboratory (8, 25, 28, 37; H. Kulaga, P.-E. Anderson, M. C. Cain, and P. J. Stopa, Proc. 1997 erdec Sci Conf. Chem. Biol. Defense Res. 1997) or the field (1, 30).Philips et al. have shown that B. anthracis spores can be identified by fluorescence microscopy and flow cytometry after appropriate labeling of the sample using fluorescent labeled antibodies (24,26,28). Nevertheless, difficulties in the selective identification of B. anthracis in mixed population samples were previously encountered (27). Recently, Stopa (37) has shown that ...

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Monoclonal antibodies against spore antigens of Bacillus anthracis

Cited by 8 publications

References 21 publications

Gamma Irradiation Can Be Used To Inactivate Bacillus anthracis Spores without Compromising the Sensitivity of Diagnostic Assays

Gamma Irradiation Can Be Used To Inactivate Bacillus anthracis Spores without Compromising the Sensitivity of Diagnostic Assays

Flow-cytometric Analysis of Bacillus anthracis Spores

Detection of Frequency Resonance Energy Transfer Pair on Double-Labeled Microsphere and Bacillus anthracis Spores by Flow Cytometry

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