The introduction of polymerase chain reaction (PCR)-based DNA amplification in the early nineties has revolutionized diagnosis of infectious diseases and has provided many fundamental insights regarding their epidemiology and clinical presentation. Compared to serology, PCR-based diagnosis may be more specific and sensitive, and may allow earlier diagnosis. Adequate patient specimens are important. Depending on the clinical setting, PCR amplification can be performed on blood samples, other clinical specimens or directly on histopathologic specimens. PCR detection of pathogen DNA is used to confirm serologic screening tests and to diagnose infections due to pathogens that are difficult to grow in culture. With chronically persistent agents, PCR may allow differentiation between clinically irrelevant and relevant infection. Suppression of viral DNA or RNA is a standard read-out of the efficacy of antiviral agents used to treat chronic hepatitis B, C, or HIV infection. Infectious disease consultation may be useful for interpretation of results. Genetic testing is increasingly being performed for antimicrobial susceptibility testing (MRSA, antitubercular treatment) and may in the future contribute to prediction of the host response to infectious disease and of antiretroviral toxicity. Increased future use of PCR-based diagnosis of infectious diseases is likely, based on increasingly automated and multiplex PCR technology.