Alpha2-macroglobulin (alpha2M) functions as an extracellular carrier of diverse cytokines, including transforming growth factor-beta1 (TGF-beta1), that expresses anti-inflammatory activities. The results presented here demonstrate that interleukin-10 (IL-10) and IL-4, which also regulate the inflammatory response, bind to alpha2M. Unlike TGF-beta, IL-4 and IL-10 bind almost exclusively to the receptor-recognized, or activated, form of alpha2M. Purified IL-4-alpha2M complexes were predominantly covalent due to thiol disulfide exchange involving Cys949 in the alpha2M subunit. Blocking Cys949 with iodoacetamide significantly inhibited IL-4- and IL-10 binding. Bovine serum albumin (BSA), which possesses a free Cys residue and undergoes thiol disulfide exchange reactions, did not compete with alpha2M for the binding of IL-4 or IL-10. These results suggest a model in which IL-4 and IL-10 associate with activated alpha2M to form complexes that are initially noncovalent but unstable. In these complexes, Cys949 is properly aligned to undergo thiol disulfide exchange and generate stable, covalent IL-4-alpha2M and IL-10-alpha2M complexes.