Using a monoclonal antibody raised against human platelet thrombospondin, we found anti‐thrombospondin immunoreactivity in the extracellular matrix of avian embryos, coincident with the ventral pathways followed by trunk neural crest cells. To confirm that the antibody recognized thrombospondin‐1 and to determine the tissue of origin of the thrombospondin matrix, a thrombospondin‐1 cRNA probe was used for whole mount in situ hybridization. This probe revealed thrombospondin‐1 mRNAs in the developing myotome before and during neural crest cell migration. The effect of thrombospondin‐1 on neural crest cell migration, morphology, and adhesion was assayed in vitro. Quail trunk neural crest cells cultured on 4 μg/ml of thrombospondin‐1 migrate at 1.14 ∓ 0.54 μm/min, which is significantly greater than the rate of cell migration on tissue culture plastic. Using a shaker‐based adhesion assay, a significantly greater number of neural crest cells remain attached to dishes coated with 4 μg/ml of thrombospondin‐1 than to tissue culture plastic alone. The number of neural crest cells that remain attached to 4 μg/ml of thrombospondin‐1 is similar to the number that remain attached to dishes coated with 10 μg/ml of fibronectin. These observations indicate that neural crest cells migrate through a thrombospondin‐filled extracellular matrix, and that thrombospondin‐1 promotes neural crest cell migration and adhesion. Thus, thrombospondin‐1 is the first somite‐derived extracellular matrix molecule with properties consistent with a role in the promotion of migration into the anterior somite, as opposed to the repulsion of neural crest cells from the posterior half of the somite. Dev Dyn 1999;214:312–322. © 1999 Wiley‐Liss, Inc.