α1-Acid glycoprotein (AGP) is a lipocalin that is produced mainly by the liver and secreted into plasma in response to infections and injuries. In this study, we evaluated AGP isoforms that can be detected in bovine milk. We found that milk-AGP content is made up of at least two isoform groups, a low MW group (44 kDa) that is produced in the mammary gland (MG-AGP), and a higher MW group (55–70 kDa), that is produced by somatic cells (SC-AGP). Identical SC-AGP isoforms can be found both in milk and blood PMN cells. Analysis of the mammary tissue cDNA showed that the sequence of the MG-AGP isoform is identical to that of plasma AGP. Each group contains several proteins with different MWs and different isoelectric points, as shown by 2D-electrophoresis. The glycosylation patterns of these isoforms were analysed by means of specific lectin binding, to evaluate the degree of sialylation, fucosylation and branching. The MG-AGP glycan pattern was identical to plasma AGP produced by the liver. Several differences were detected, however, between plasma and SC-AGP isoforms, the most evident being the strong degree of fucosylation and the elevated number of di-antennary glycans in SC-AGP. Immunohistochemistry showed that AGP is found in all tissues that make up the mammary gland, but that it is most likely produced for the main part by the alveoli.