The detection and characterization of microbial agents in biological specimens are essential for the investigation of disease outbreaks, for epidemiologic studies of the clinical course of infections, and for the assessment of the role of infectious agents in chronic diseases. Methodological approaches depend on the infectious agent, the specimens analysed and the target populations. Although the diagnosis of infectious diseases has traditionally relied on direct microscopic examination of samples and on the cultivation of microbial agents in vitro, novel techniques with increased sensitivity and specificity are now being used on samples that can be more easily collected and transported to microbiology laboratories (e.g. dried blood spots on filter paper for nucleic acid analysis). Direct detection techniques include the microscopic examination of specimens with special stains, antigen detection and nucleic acid detection by molecular assays. These assays are highly sensitive and provide rapid results for most agents. Genomic amplification assays greatly increase the sensitivity of nucleic acid-based tests by extensive amplification of specific nucleic acid sequences before detection. Real-time polymerase chain reaction (PCR) permits genomic amplification concurrently with detection of amplified products. Typing infectious agents requires additional investigation employing either serologic techniques to identify unique antigenic epitopes, or molecular techniques. These studies are important for epidemiologic purposes, as well as for the investigation of pathogenesis, disease progression, and to establish causality between a disease and a microbial agent. Much of bacteriology has relied on growth of organisms on artificial media, and on identification of bacterial growth with biochemical, serological, or more recently, nucleic acid-based tests. The detection of specific antibodies from the host directed against pathogens is another strategy to identify current or past infections.