Polyamidoamine (PAMAM) dendrimer/DNA complexes encapsulated in a water soluble polymer, poly-alpha,beta-[N-(2-hydroxyethyl)-l-aspartamide], were supported on a cholic acid functionalized star poly(DL-lactide) film with a fast degradation rate to mediate localized gene delivery. The in vitro gene transfections of two types of cells, HEK293 and NIH3T3, were investigated. The expressions of pGL3-Luc and pEGFP-C1 plasmids in HEK293 cells indicated that the star poly(DL-lactide) supported PHEA encapsulated PAMAM/DNA complexes could effectively mediate transfection, with transfection efficiencies which were comparable to that of solution-based transfections. Whereas the PAMAM/DNA complexes directly supported on the star poly(DL-lactide) film showed a much lower expression level for HEK293, which indicated the existence of PHEA played an important role in the efficient transfection. The solid support-based transfection for NIH3T3 cells exhibited higher expressions of pGL3-Luc compared with the solution-based transfection. Encapsulating PAMAM/DNA complexes in PHEA could further improve the gene expression in NIH3T3. During the cellular transfection, the degradation of the cholic acid functionalized star poly(DL-lactide) film could be obviously detected and the degradation did not show any unfavorable effects on the gene expression, which implied this solid support-based gene delivery device had great potential for localized transfection.