Transforming growth factor β (TGF-β) is one of the predominant growt h factors present in milk. The concentration, molecular mass forms and stability of TGF-β in bovine milk were investigated using a standard bioassay measuring the growth inhibition of a mink lung epithelial cell line. Most of the TGF-β bioactivity in milk was found to be in a latent form, which was also retained in the whey fraction. After acid activation, the total TGF-β concentration was 4.3 ± 0.8 ng and 3.7 ± 0.7 ng TGF-β per ml of milk and cheese whey respectively. Cation-exchange chromatography at pH 6.5 was used to concentrate latent whey-derived TGF-β, which could be activated by transient exposure to extremes of pH, urea or heat. Heparin did not significantly activate milk-derived TGF-β. Neutral gel filtration of the cationic whey fraction revealed a major peak of latent TGF-β with a molecular mass of 80 kDa and a smaller peak at 600 kDa. Transient acidification of the cationic whey fraction prior to neutral gel filtration, or gel filtration under acidic conditions, released low molecular mass TGF-β from both high molecular mass peaks. Whey-derived TGF-β was purified using a five-step chromatographic procedure. An N-terminal sequence was obtained for TGF-β2, which accounted for over 85% of the TGF-β bioactivity in whey. All TGF-β activity in whey could be neutralised by a monoclonal antibody directed against TGF-β1, -β2 and -β3. The results suggest that the majority of TGF-β in bovine milk is present in a small latent complex.