Here we describe the isolation and identification of a major cartilage glycoprotein which is co-extracted along with typical hyaline cartilage components such as collagen types II and IX from chicken embryo sternum. In polyacrylamide gel electrophoresis it migrates as a high molecular mass protein (greater than 10(6) daltons) which on reduction gives rise to a prominent doublet at 205/195 kd and minor bands at 220 and 170 kd. The intact molecule sediments as a 13S component in rate zonal centrifugation, indicative of a highly extended conformation in solution. In these properties it closely resembles myotendinous antigen, a glycoprotein recently detected in a number of embryonic tissues, including cartilage. This identity was confirmed by immunoblotting using a monoclonal antibody (M1) specific for myotendinous antigen. Electron micrographs of the rotary shadowed molecule revealed an unusual six-armed structure, indistinguishable in form and dimensions from hexabrachion, a recently discovered contaminant of cellular fibronectin preparations. These structures could be decorated with the M1-antibody, demonstrating that hexabrachion is myotendinous antigen. This extended, potentially multivalent molecule could provide an ideal substrate to connect widely spaced components in a highly hydrated tissue such as cartilage.