More and more severe infections with Clostridium difficile (Cd) are occurring worldwide. This increase in infections and infections with more virulent (Cd) highlight the need for a rapid and sensitive diagnostic assay for the presence of Cd.  A laboratory derived PCR described here identifies common hospital strains as well as more recently described genetic variants. We compared our laboratory real time PCR assay for Cd Toxin B to both B-D GeneOhm Toxin B commercial PCR assay and Wampole^TM C DIFF QUIK CHEK COMPLETE EIA assay. The laboratory derived PCR compared to the B-D GeneOhm Toxin B commercial PCR assay had a sensitivity of 98.2%, sensitivity of 100%, positive predictive value of 100% and negative predictive value of 95%. The laboratory derived assay is more sensitive than Wampole^TM  C DIFF QUIK CHEK COMPLETE EIA assay. The laboratory derived PCR also has lower cost than commercially available PCR and EIA assays. Its use on all stools eliminates the need to resolve discrepant EIA results and shortens over-all reporting times.