Stanniocalcin-1 (STC-1) is a novel calcium- and phosphate-regulating hormone that was originally found in bony fish and subsequently in mammals. This study mainly aimed to outline the distribution and relative expression levels of STC-1 in the gastrointestinal tract and kidney of neonatal Holstein calves. The localization of the STC-1 protein/mRNA in these sites was investigated by immunocytochemistry and in situ hybridization staining. Impressively, the results showed that the STC-1 mRNA was mainly distributed in the distal segments of the kidney, whereas the protein was mostly found in the proximal and distal segments, including the glomerulus. In the gastrointestinal tract, the STC-1 mRNA was expressed in the lamina propria and mucosa epithelial cells, whereas the protein was found in the cytoplasm of goblet cells, chief cells of the fundic glands, gastric pits, and intestinal glands. Quantitative analyses indicated that the highest levels of both STC-1 mRNA and protein were expressed in the kidney. The highest STC-1 mRNA and protein levels in the stomach were expressed in the abomasum. In addition, the duodenum and colon exhibited the highest STC-1 mRNA relative expression ratios among the intestine, whereas the highest protein expression levels were found in the duodenum and jejunum. In addition, The STC-1 protein detected in this experiment may be a STC50 rather than a big STC. Thus, STC-1 may be involved in digestion and/or absorption in the alimentary canals and the kidney in those sites with active digestion and absorption.