Nucleic acid technology has assumed an essential role in various areas of in vitro diagnosis. Its major applications include the genomic characterization of mutations and polymorphisms, amplification of nucleic acids by the polymerase chain reaction, analysis of gene expression patterns at the mRNA level, specific detection of mutant proteins, and engineering of proteins used in ligand binding assay. Manipulating the expression of genes in cells and experimental animals allows detailed analysis of processes leading from genomic alterations to disease manifestations and may ultimately yield refinements in diagnostic strategies. The addition of DNA and RNA to the conventional diagnostic in vitro targets and novel approaches to the study of disease processes and manifestations characterize a diagnostic strategy referred to as "molecular diagnostics." This contribution describes some of essential ways in which molecular diagnostics differs from conventional diagnostic approaches. Applications such as differential diagnosis, prenatal diagnosis, early diagnosis, and diagnosis of disease susceptibility are well established in single-gene disorders, and the diagnostic impact of DNA polymorphisms is steadily increasing in multifactorial diseases. The high sensitivity of the polymerase chain reaction makes possible the detection of single infectious agents or tumor cells. With increasing knowledge of expressed gene sequences the expression pattern of mRNA will reflect the biological state of cells with high precision. Mutant proteins can be analyzed based on their structural or biological properties. Use of the appropriate expression systems makes it possible to design proteins for diagnostic in vitro applications such as ligand binding assays.