Development of a rapid and sensitive immunomagnetic-bead based assay for detecting Bacillus cereus in milk

摘要

Bacillus cereus is a major food-born pathogen in Taiwan and its major syndromes include vomiting, fever and diarrhea. To minimize the possibility of exposing consumers to pathogenic B. cereus, this study develops a rapid and sensitive assay that utilizes immunoliposomal nanovesicles (IMLNs) and immunomagnetic beads (IMBs). In this work, fluorescent dyes (sulforhodamine B)-loaded IMLNs were employed to increase the detection signal; anti-B. cereus antibody-conjugated IMBs were applied to capture B. cereus in samples. Hence in this assay, a sandwich complex was formed as “IMBs-B. cereus-IMLNs”. The optimal IMLNs had a diameter of 300 nm with a conjugated antibody molar percentage (mol%) of 0.25 mol%. The limit of detection (LOD) of this developed assay reaches 10 CFU/mL of B. cereus with the false negative value as zero in 20 parallel assays in milk samples. To evaluate the specificity of this assay, nine Gram positive and negative bacteria were tested and found to cause no significant interference problems. In conclusion, this study elucidates the feasibility of using a novel IMB/IMLN assay for detecting B. cereus and its LOD without pre-enrichment could amount to 10 CFU/mL within 4 h.