No abstract

Background: Recent use of Bacillus anthracis spores as a bioweapon has highlighted the need for a continuous monitoring system. Current monitoring systems rely on antibody-derived probes, which are not hardy enough to withstand long-term use under extreme conditions. We describe new, phage-derived probes that can be used as robust substitutes for antibodies. Methods: From a landscape phage library with random octapeptides displayed on all copies of the major phage coat protein of the phage fd-tet, we selected clones that bound to spores of B. anthracis (Sterne strain). ELISA, micropanning, and coprecipitation assays were used to evaluate the specificity and selectivity with which these phage bound to B. anthracis spores. Results: Peptides on the selected clones directed binding of the phage to B. anthracis spores. Most clones exhibited little or no binding to spores of distantly related Bacillus species, but some binding was observed with spores of closely related species. Our most specific spore-binding phage displayed a peptide EPRLSPHS (several thousand peptides per phage) and bound 3.5-to 70-fold better to spores of B. anthracis Sterne than to spores of other Bacillus species. Conclusions: The selected phage probes bound preferentially to B. anthracis Sterne spores compared with other Bacillus species. These phage could possibly be further developed into highly specific and robust

No abstract

No abstract

Recent outbreaks of food borne illnesses continue to support the need for rapid and sensitive methods for detection of foodborne pathogens. A method for detecting Listeria monocytogenes in food samples was developed using an automated fiber-optic-based immunosensor, RAPTOR ™. Detection of L. monocytogenes in phosphate buffered saline (PBS) was performed to evaluate both static and flow through antibody immobilization methods for capture antibodies in a sandwich assay. Subsequent detection in frankfurter samples was conducted using a flow through immobilization system. A two stage blocking using biotinylated bovine serum albumin (b-BSA) and BSA was effectively employed to reduce the non-specific binding. The sandwich assay using static or flow through mode of antibody immobilization could detect 1×10 3 cfu/ml in PBS. However, the effective disassociation constant K d and the binding valences for static modes of antibody immobilization in spiked PBS samples was 4×10 5 cfu/ml and 4.9 as compared to 7×10 4 cfu/ml and 3.9 for flow through method of antibody immobilization. Thus the sensitive flow-through immobilization method was used to test food samples, which could detect 5×10 5 cfu/ml of L. monocytogenes in frankfurter sample. The responses at the lowest detectable cell numbers in the frankfurter samples was 92.5 ± 14.6 pA for L. monocytogenes to comparative responses of 27.9 ± 12.2 and 31 ± 14.04 pA obtained from Enterococcus Sensors 2006, 6 809 faecalis and Lactobacillus rhamnosus (control species), respectively. The effective K d and binding valency from spiked frankfurter samples was 4.8×10 5 cfu/ml and 3.1, thus showing highly sensitive detection can be achieved using the RAPTOR ™ biosensor even in the presence of other bacterial species in the matrix.