We report on the synthesis of well-defined amphiphilic copolymer brushes possessing alternating poly(methyl methacrylate) and poly(N-isopropylacrylamide) grafts, poly(PMMA-alt-PNIPAM), via a combination of atom transfer radical polymerization (ATRP) and click reaction (Scheme 1). Firstly, the alternating copolymerization of N-[2-(2-bromoisobutyryloxy)ethyl]maleimide (BIBEMI) with 4-vinylbenzyl azide (VBA) affords poly(BIBEMI-alt-VBA). Bearing bromine and azide moieties arranged in an alternating manner, multifunctional poly(BIBEMI-alt-VBA) is capable of initiating ATRP and participating in click reaction. The subsequent ATRP of methyl methacrylate (MMA) using poly(BIBEMI-alt-VBA) as the macroinitiator leads to poly(PMMA-alt-VBA) copolymer brush. Finally, amphiphilic poly(PMMA-alt-PNIPAM) copolymer brush bearing alternating PMMA and PNIPAM grafts is synthesized via the click reaction of poly(PMMA-alt-VBA) with an excess of alkynyl-terminated PNIPAM (alkynyl-PNIPAM). The click coupling efficiency of PNIPAM grafts is determined to be similar to 80%. Differential scanning calorimetry (DSC) analysis of poly (PMMA-alt-PNIPAM) reveals two glass transition temperatures (T-g). In aqueous solution, poly(PMMA-alt-PNIPAM) supramolecularly selfassembles into spherical micelles consisting of PMMA cores and thermoresponsive PNIPAM coronas, which were characterized via a combination of temperature-dependent optical transmittance, micro-differential scanning calorimetry (micro-DSC), dynamic and static laser light scattering (LLS), and transmission electron microscopy (TEM). (C) 2009 Wiley Periodicals, Inc. J Polym Sci Part A: Polym Chem 47: 2608-2619, 2009