A Colony Blot Immunoassay for the Rapid Identification of Bacillus cereus

Abstract

A colony blot immunoassay was developed for the rapid identification of Bacillus cereus using antibodies against the 28.5-kDa cell-surface antigen of B. cereus. Suspect colonies from plates were blotted onto a Whatman #541 membrane, dried, and fixed by UV irradiation. The membrane was then immersed in an anti- B. cereus antibody-horseradish peroxidase conjugate for 60 min. After washing and reacting with 4-chloro-1-naphthol and H2O2, the appearance of purple spots indicated the presence of B. cereus. This assay effectively identified 61 of 62 B. cereus strains tested. Among 38 non– B. cereus strains, which were other Bacillus spp. (19 genera), 36 gave true-negative results, and 2 showed false-positive results. The sensitivity and specificity for B. cereus were 98.4 and 94.7%, respectively. The present assay is easy to use, and the rapid identification of B. cereus can be completed in 2.5 h.