In this study, the cytotoxicity and transfection efficiency of using chitosan/DNA complex combined with poly(ethylenimine) (PEI) were investigated. The combination of PEI with the chitosan/DNA complex markedly enhanced the gene expression of HeLa cells to 1000-fold of that induced by chitosan alone. PEI's cytotoxicity was considerably decreased upon combination with the chitosan/DNA complex. Furthermore, the PEI/chitosan/DNA could maintain the gene expression efficiency in the presence of serum.