In this study, it is investigated the elimination of residual sodium ions from sodium silicate (Na2SiO3) modified Fe3O4@SiO2 nanoparticles, via the additional acidic treatment. Residual sodium ions are generated from the Na2SiO3 precursor during the titration for the fabrication of the Fe3O4@SiO2 core-shell structure; however, they were entirely removed after the in situ additional titration at pH 3 using a 1 M of hydrochloric acid (HCl). The thickness of the SiO2 shell layer of each nanoparticle was measured to 10 nm and the morphology of SiO2 shell on the Fe3O4 core were not significantly different after acid treatment to remove the residual sodium ions. However, the absolute zeta potential of the Na2SiO3-modified Fe3O4@SiO2 was decreased about 20% after HCl treatment (−52.5 ± 3.0 mV). In addition, the mean particle size of the Na2SiO3-modified Fe3O4@SiO2 was increased (363.98 nm) and the polydispersity index, which indicates dispersibility was increased (0.23) since residual sodium ions were eliminated by the acid treatment. It is assumed that the reducing anions in the diffuse layer of the particle due to the elimination of the residual sodium, as the counter ion, decreased the surface charge, thereby reducing the repulsive force between the Fe3O4@SiO2 nanoparticles. Then Fe3O4@SiO2 nanoparticles were utilized for separating plasmid DNA and separated plasmid DNA was measured with agarose gel electrophoresis and the absorbance monitoring. The efficiency of the DNA purification of the Na2SiO3-modified Fe3O4@SiO2 measured to 43.3 ± 5.2 ng/μl which was 58% increased after acid treatment. As a result, the dispersion/magnetic properties and DNA purification efficiency of the Na2SiO3-modified Fe3O4 became equivalent to the characteristics of the TEOS-modified Fe3O4@SiO2 using the additional acid treatment for sodium ions removal.