抄録
Bovine β-lactoglobulin (β-LG) was conjugated with chitosan (CHS) by means of a water-soluble carbodiimide to reduce the immunogenicity of β-LG. Each β-LG-CHS conjugate was purified by ion-exchange chromatography and hydrophobic chromatography. The conjugation between β-LG and CHS was confirmed by SDS–PAGE, the isoelectric point of the conjugate being higher than that of β-LG. Two types of the β-LG-CHS conjugate were obtained with molar ratios of β-LG to CHS of 1:1 (F1) and 1:2 (F2). Structural analyses by fluorescence measurement, ELISA with monoclonal antibodies and retinol-binding activity indicated that the conjugates had almost maintained the native structure of β-LG. The antigenicity of the β-LG-CHS conjugates was similar to that of β-LG in C3H/He mice. Reduction of the immunogemicity of β-LG was achieved by conjugation with CHS. In particular, F2 showed very low immunogenicity. B cell epitopes of β-LG and the conjugates recognized in C3H/He mice were determined with 15-mer multi-pin peptide; the linear epitope profiles of the conjugates were found to be similar to those of β-LG, while the antibody response to each epitope was dramatically reduced. Conjugation of β-LG with chitosan was effective for reducing the immunogenicity of β-LG.
