Thermosensitive copolymer of N-vinylcaprolactam (VCL) and 1-vinylimidazole (VI), a new reagent for metal-chelate affinity separation of proteins, was characterized by static and dynamic light scattering and H-1 NMR spectrometry. The copolymer of M-w = 11 700, M-w/M-n 1.1 and VI-unit molar fraction of 11% was prepared by free radical co-polymerization of the monomers followed by dialysis of the product. The copolymer solution in 10 mM Na-phosphate buffer containing 0.15 M NaCl, pH 8.5, at 25 degreesC turned out to be close to theta -conditions. Its diffusion coefficient in these conditions was 6 x 10(-7) cm(2) s(-1) indicating the hydrodynamic diameter of 8 nm. The copolymer was separated by immobilized metal affinity chromatography on Cu2+-IDA-Sepharose into the non-adsorbing fraction (5 molar% VI-units) and the adsorbing fraction further eluted with EDTA and thermally precipitated at 48 degreesC (8.5 mol% VI-units). The combination of IMAC and thermal precipitation of the copolymer enables isolation of its high-affinity, relatively homogeneous fraction most promising for bipseparations, including displacement chromatography of proteins. (C) 2001 Elsevier Science Ltd. All rights reserved.