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 probes suitable for long-term use in continuous monitoring devices and biosorbents.