Clostridium difficile is an anaerobic, gram-positive bacillus that is the leading cause of antibiotic associated diarrhea and pseudomembranous colitis in health care facilities. Incidence of Clostridium difficile infection (CDI) has been increasing, and severe cases are becoming more common. CDI disease symptoms range from mild diarrhea to severe colitis, and even bowel perforation and death. The most common risk factor is exposure to antibiotics. The diagnosis of Clostridium difficile infection is based upon the clinical signs and symptoms, such as diarrhea, and laboratory or pathologic tests. Laboratory diagnosis of toxigenic Clostridium difficile includes anaerobic culture followed by detection of toxin or detection of toxin gene(s) or enzyme(s) found in the stool. It appears that toxin B is necessary for the development of CDI. Recently, PCR methods for the detection of toxin A and/or toxin B have been developed with high sensitivity and specificity as compared to the cell cytotoxicity and immunoassays. The combination of these characteristics may allow for prompt targeted treatment of CDI patients and thus potential improvement in patient outcome, reduced recovery time, and infection control practices.