The colony lift immunoassay (CLI), a novel method for rapid immunodetection and enumeration of target pathogens following specimen preparation, has been applied to enterohemorrhagic E. coli O157:H7. The CLI, when used in conjunction with a selective and differential medium, enables the detection, confirmation, and quantification of E. coli O157:H7 organisms after the colonies are isolated onto agar or filter monitor membranes with high sensitivity (100%) and specificity (88.9%), based on the data derived from this study (Table 1).