In this study, ABC-type triblock copolymers, poly(ethylene glycol)-b-poly(epsilon-caprolactone)-b-poly(2-(2-aminoethyl amino) ethyl methacrylate)s (PEG-PCL-PAEAEMAs), composed of novel poly(2-(2-aminoethyl amino) ethyl methacrylate) (PAEAEMA) that have a primary and a secondary amino group in each monomeric unit, were synthesized successfully. It was found that the length of PAEAEMA segments did not have obvious influence on the DNA-binding capacity and other biophysical properties (size and zeta potential values of polymer/pDNA complexes), but longer PAEAEMA chains led to a better buffering capacity. The triblock copolymers could mediate efficient gene expression that was similar to branched 25-kDa polyethylenimine (25 kDa PEI) in the absence of serum and even superior to 25 kDa PEI in the presence of serum in COS-7 cells. Low cytotoxicity of these polymers was also found in COS-7 cells. As a result, PEG-PCL-PAEAEMAs are attractive candidates as serum-tolerable gene carriers in biomedical field. (C) 2011 Wiley Periodicals, Inc. J Appl Polym Sci 121: 666-674, 2011