"Stat" multiplex polymerase chain reaction: its role in clinical microbiology.

In the laboratory, the department most affected by the introduction of molecular diagnostic tools has been the clinical microbiology laboratory. For organisms such as Clostridium difficile, polymerase chain reaction (PCR) is the gold standard for diagnosis. Incorporation of molecular diagnostic techniques into microbiology laboratories of differing sizes has been hastened by the development of simplified technological methods, individual cassettes incorporating all reagents necessary for diagnosis, and instruments allowing multiple tests to be run at the same time. Companies, such as Cepheid (Sunnyvale, CA), with its GeneXpert technology , even advertise that their tests can be run in a physician's office. Pressure by the US Food and Drug Administration (FDA) on companies such as Roche has pushed them to change their offerings in molecular diagnosis from analyte-specific reagent kits to a commercially prepared, prepackaged assay with detailed instructions for performance of the test. This change has hastened adoption of molecular methods into the testing menu of laboratories that do not have the resources and staff to perform the extensive and time-consuming validations of assays that are not approved by the FDA. Molecular methods have probably found their greatest use in microbiology laboratories in the diagnosis of viral infections. The reasons for the movement to molecular methods in this arena are varied. The rapid assays for agents such as influenza A and B and respiratory syncytial virus have low sensitivity and specificity. Direct fluorescent antibody assays require trained technologists and are generally only available during the daytime and early evening hours. Culture is less sensitive than molecular assays. The main advantage of molecular assays over conventional methods is that they can detect newer viruses that are often unculturable. Multiplex assays that detect multiple organisms in a single procedure are joining singleplex tests that detect nucleic acids from a single species. However, even the newer multiplex PCR tests, which have high sensitivity and specificity, are complicated, are technically demanding, and require highly specialized technologists. The turnaround time for these assays is fairly long (ie, 6-8 hours). As a result, these tests are generally only performed during daytime hours in virology/molecular laboratories in large, tertiary medical centers. In this issue of the Journal, Xu and colleagues 1 have demonstrated the advantages of using a multiplex PCR assay to diagnose respiratory viral infections in a core laboratory serving a patient population presenting to the emergency department. Performance of the panel required a hands-on time of …