A carboxymethyl poly(1-vinylimidazole) (CM-PVIm) has been synthesized as a polyampholyte for biocompatibility. A solution turbidity measurement proved that no significant effect on rapid aggregate formation or phase separation of serum proteins is induced by the resulting polyampholyte CM-PVIm. Agarose gel retardation assay shows no significant interaction of the CM-PVIm with DNA. However, zeta-potential measurement proved that the CM-PVIm coated poly(ethylenimine) (PEI)/DNA binary complexes. The resulting CM-PVIm/PEI/DNA ternary complexes largely preserved the gene expression mediated by the PEI/DNA binary complexes with the cell viability increased. These results suggest that the CM-PVIm can be a polyampholyte for biocompatibility.