Three isozymes of nitric oxide synthase (NOS) have been identified, cDNAs isolated and sequenced, and antibodies produced against each isozyme. Isozyme I (found primarily in central and peripheral neuronal cells), II (in cytokine-induced cells), and III (in endothelial cells) show less than 58% identity in the deduced amino acid sequences from humans. Many investigators have produced isozyme-specific antibodies and used these antibodies to locate these proteins in various cells and tissues. NOS-I is constitutively expressed, and the enzymatic activity is regulated by Ca2+ and calmodulin. The anti-NOS-I antibodies have allowed investigators to characterize non-adrenergic non-cholinergic neurons as nitrergic neurons, revealed NOS-I immunoreactivity in neurons and macula densa cells of the kidney and pancreatic islet cells, human skeletal muscle, and to demonstrate that various structures within the brain and spinal cord contain NOS-I. NOS-II is not regulated by Ca2+ and has been implicated in the pathophysiology of sepsis and autoimmune diseases. The anti-NOS-II antibodies have localized this isoform to infiltrating macrophages in pancreatic islets of diabetic rats, infiltrating macrophages and myocytes of a transplant heart model in rats, various cell types in bacterially and endotoxin-treated rats, alveolar macrophages in areas of inflammation in humans, and vascular smooth muscle cells of human atherosclerotic aneurysm. Isoform III is similar to NOS-I in that it is constitutively expressed and regulated by Ca2+ and calmodulin. Anti-NOS-III antibodies have found that this isoform is relatively specific for endothelial cells.