The novel real-time PCR assay developed as described here was able to detect bla(KPC1/2-12) (bla(KPC-1/2) to bla(KPC-12)) from easily available clinical specimens in less than 2 h. The genotypic assay was highly sensitive (100%) and specific (98%). In some cases, it was able to detect bla(KPC) 48 h before positive detection by standard phenotypic assay on patients who were monitored daily. The high sensitivity and rapidity of the molecular method make it the method of choice for KPC surveillance and, thus, containment purposes.