The introduction of molecular diagnostic methods in the microbiologic diagnostic laboratory is subject to a number of practical and financial constraints requiring the elaboration of a sound strategy. Validation of the tests may require the use of an expanded gold standard or the application of novel statistical methods such as latent class analysis. The procedures should be constantly evaluated for false positive an false negative results. The introduction of molecular methods will not only depend on their performance for each individual microorganism, but also on the clinical relevance of the diagnostic question asked, the prevalence of the clinical problem and whether the new methods are added to the procedures in use or will replace them. Therefore no general rules can be proposed, strategies have to be elaborated for each infectious agent or clinical syndrome. Strategies concerning the use of molecular diagnostic techniques for the diagnosis of Mycobacterium tuberculosis, Chlamydia trachomatis, meningo-encephalitis syndrome and respiratory infections, are discussed. Their role in typing of microorganisms and the detection of previously unknown infectious agents are mentioned.