The purpose of this study was to purify soybean beta-conglycinin subunits and analyze their IgE-binding properties with IgE immunoblotting and their response in a basophil histamine release test. The method presented in this paper offers an effective procedure to purify beta-conglycinin subunits with higher yield and purity compared with previous methods by using a combination of ion exchange and metal chelate affinity chromatography. Beginning with a column load of 600 mg beta-conglycinin, the new method yielded 100.4 mg of protein containing 96.7% alpha subunit, 97.0 mg of protein containing 91.6% alpha' subunit and 89.4 mg of protein containing 96.1% beta subunit. All of the purified subunits of beta-conglycinin reacted with IgE from sera of soybean-allergic patients and induced dose-dependent histamine release from basophils from these patients, suggesting that purified alpha, alpha', and beta subunits of beta-conglycinin retained allergenic activity and could be used for in vitro and in vivo diagnosis of soybean-induced food allergy. (C) 2012 Published by Elsevier Ltd.