Controversy remains regarding the origin of the pancreatic endocrine cells. It is generally accepted that the majority of insulin-secreting cells derive from the endodermal epithelium of the gastrointestinal tract. The aim of this study was to determine the contribution made by a particular cluster of differentiation (CD)-positive cells to the development of the bovine endocrine pancreas. In bovine embryos and foetuses with crown to rump lengths (CRL) ranging from 1 to 47 cm, cells staining positively for CD34 and/or CD133 were always more numerous in the left lobe and body of pancreas than in the right lobe. In the early stages of pancreatic development (CRL <5 cm), CD34 and/or CD133-reactive cells were concentrated within the epithelial cell cords that form the primitive pancreas. In later developmental stages (CRL >5 cm), individual or groups of CD34 and/or CD133-reactive cells were present in newly formed acini, which bulged out from the duct system that had arisen from the cords. Some of the positively stained cells accumulated in focal areas associated with hyperplastic intra-acinar cells. These "acino-insula-like complexes" appeared to enlarge with age and develop into intralobular Islets of Langerhans. Most of the described CD34 and/or CD133-reactive cells displayed co-localisation with glucagon. A negligible number of these cells showed co-localisation with insulin. Glucagon-stained cells were distinct from insulin-stained cells and were more abundant in embryonic and early foetal pancreata. Our data demonstrate that CD34 and/or CD133-reactive cells contribute to the pancreatic alpha cell population during early foetal development in cattle.