Novel films based on cross-linked hyperbranched poly(amine-ester) (HPAE) were prepared by cross-linking the terminal hydroxyl groups of HPAE using glutaraldehyde (GA). The cross-linking process was monitored by measuring the intrinsic viscosity of HPAE/GA in N,N-dimethylacetamide. The surface structures of the cross-linked HPAE films obtained from different HPAE/GA ratios were imaged using atom force microscopy, and their properties were characterized in terms of hydrophilicity, solvent swelling, mechanics, and protein adsorption. It was found that the static contact angle was <32.9 • , tensile strength was >0.35 MPa, elongation at break was >9.2 %, swelling degree was >63 % in water, and bovine serum albumin adsorption was relatively low. The results indicate that cross-linked HPAE films have a strong application potential in many areas.