## 3 Results and Discussion  
First of all, we examined the effects of superparamagnetic nanoparticles on PCR amplification without an external magnetic field. Amplification was then performed while changing the concentration of the superparamagnetic nanoparticles mixed with the PCR reagent. Figure 2(a-c) show amplified DNA bands with different concentrations of magnetic nanoparticles observed using agarose gel electrophoresis. Each corresponds to the inset photograph of diluted magnetic nanoparticles with the same concentration. With EMG607 magnetic nanoparticles, PCR amplification was performedwith particle concentrations of 0.4-1.8 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L. It was observed that the fluorescence intensity of DNA bands decreased with higher levels of nanoparticle concentration, and consequently PCR amplification was inhibited at higher concentrations, as shown in Figure 2(a). With EMG707 magnetic nanoparticles, PCR amplification was performed with particle concentrations of 0.04-0.18 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L, and further amplification using EMG1111 magnetic nanoparticles was carried out with particle concentrations of 0.02-0.12 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L. Magnetic nanoparticles of the EMG707 and EMG1111 types also inhibited PCR amplification at high concentrations, as shown in Figure 2(b,c). Figure 2(d) shows the dependence of product yield on the concentration of the magnetic nanoparticles added to PCR reagents in quantitative terms. It clearly indicates that the yield gradually decreased with higher levels of concentration, and as a result, PCR amplification was completely inhibited at threshold concentrations. The same figure also shows that the threshold concentrations for inhibited PCR amplification of EMG607, EMG707 and EMG1111 can be determined as 0.8, 0.12 and 0.08 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L, respectively, and that threshold concentration strongly depends on the particle type. Figure 2: Amplified DNA bands with different concentrations of magnetic nanoparticles observed using agarose gel electrophoresis, each corresponding to the inset photograph of diluted magnetic nanoparticles with the same concentration. (**a**) Lane 1: control without magnetic nanoparticles; lanes 2–9: 0.4, 0.6, 0.8, 1.0, 1.2, 1.4, 1.6 and 1.8 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L of EMG607 magnetic nanoparticles, respectively (**b**) Lane 1: control without magnetic nanoparticles; lanes 2–9: 0.04, 0.06, 0.08, 0.1, 0.12, 0.14, 0.16 and 0.18 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L of EMG707 magnetic nanoparticles, respectively (**c**) Lane 1: control without magnetic nanoparticles; lanes 2–9: 0.02, 0.03, 0.04, 0.05, 0.06, 0.08, 0.1 and 0.12 \(\mathrm{\SIUnitSymbolMicro\SIUnitSymbolMicro}\)L of EMG1111 magnetic nanoparticles, respectively. (**d**) Dependence of PCR product yield on the concentration of the magnetic nanoparticles added to the PCR solution. EMG707 and EMG1111 magnetic nanoparticles have a low ratio of particles to PCR reagent and inhibit PCR amplification at low concentrations, while the EMG607 type with cationic surfactant modification has a high ratio of particles to PCR reagent and becomes effective at much higher concentrations. These results indicate that magnetic nanoparticles as well as gold nanoparticles suppressed PCR efficiency remarkably, thereby inhibiting PCR amplification. Consequently, we consider that an inhibitory effect is induced by complex interaction between nanoparticles and PCR reagents depending on particle surface properties. To clarify the nature of the PCR inhibition effect, the influence of magnetic nanoparticles on specific PCR reagents was elucidated.