The paper describes the isolation and characterization of a new 350-kD glycoprotein called fibrillin, which is a component of extracellular microfibrils. Fibrillin was isolated from the medium of human fibroblast cell cultures and characterized by electrophoresis, immunoblotting, and immunofluorescence. It was found to have intrachain disulfide bonds, stain with periodic acid-Schiff reagent, and contain [³H]glucosamine but not [³S]sulfate. Fibrillin was resistant to bacterial collagenase digestion. Electron microscopic immunolocalization using colloidal gold conjugates showed that fibrillin is distributed in a class of extracellular structural elements known as microfibrils, which have an average diameter of 10 nm. These microfibrils were found around the amorphous cores of elastic fibers and in bundles without elastin cores. The periodicity of fibrillin's epitope matched the interstitial collagen band periodicity. In contrast, type VI collagen, which has been proposed as a possible microfibrillar component, was localized to small diameter microfilaments associated with large banded collagen fibers. The study suggests that fibrillin is a major structural component of microfibrils and provides insights into the organization and function of these extracellular structures.The paper describes the isolation and characterization of a new 350-kD glycoprotein called fibrillin, which is a component of extracellular microfibrils. Fibrillin was isolated from the medium of human fibroblast cell cultures and characterized by electrophoresis, immunoblotting, and immunofluorescence. It was found to have intrachain disulfide bonds, stain with periodic acid-Schiff reagent, and contain [³H]glucosamine but not [³S]sulfate. Fibrillin was resistant to bacterial collagenase digestion. Electron microscopic immunolocalization using colloidal gold conjugates showed that fibrillin is distributed in a class of extracellular structural elements known as microfibrils, which have an average diameter of 10 nm. These microfibrils were found around the amorphous cores of elastic fibers and in bundles without elastin cores. The periodicity of fibrillin's epitope matched the interstitial collagen band periodicity. In contrast, type VI collagen, which has been proposed as a possible microfibrillar component, was localized to small diameter microfilaments associated with large banded collagen fibers. The study suggests that fibrillin is a major structural component of microfibrils and provides insights into the organization and function of these extracellular structures.