## 3 Results
### AFM Study of the Effect of Pulsed Electromagnetic Field  
Figure 3 displays typical AFM images obtained upon AFM scanning of the surface of mica substrate after its incubation in working and control solutions of HRP, which were irradiated and not irradiated in the pulsed electromagnetic field, respectively.  
Figure 3: Results of AFM study of the effect of PEMF on the HRP aggregation. (**A–D**) Typical AFM images (**A,C**) and cross-section profiles (**B,D**) of the surface of mica substrate obtained after its incubation in 0.1 uM aqueous HRP solution, which was either not irradiated (**A,B**) or irradiated (**C,D**) in PEMF. The cross-section profiles correspond to the lines in the respective AFM images. (**E**) Distribution plots of the AFM-visualized objects with height _p(h)_, obtained for the PEMF-irradiated (red curve) and for the control HRP solution (black curve).  
Panel (A) displays the image obtained in the control experiment. In this experiment, no electromagnetic impact on the analyzed enzyme solution was performed. As seen from Figure 3A,B, in the case of the absence of an electromagnetic field impact on the protein solution, it adsorbs onto the substrate surface in the form of compact objects, whose height makes up 1 to 2 nm. After processing the AFM data obtained, a distribution of the visualized objects with height \(\rho\)(_h_) was plotted. The \(\rho\)(_h_) distribution plot is shown in Figure 3E (black curve). The \(\rho\)(_h_) distribution curve indicates that the maximum number of the visualized objects have a height of about (1.0 \(\pm\) 0.2) nm, while the width at half-height makes up 0.8 nm.  
The molecular weight of HRP is known to be 40 to 44 kDa [35; 36]. Moreover, in previous studies, it was demonstrated that proteins of similar molecular weight--for instance, monomers of various cytochromes P450--also have a height of about 1 to 2 nm [67; 68]. For these considerations, we have concluded that the objects with a height of 1 to 2 nm, observed during AFM scanning, can be attributed to HRP monomers. At the same time, the \(\rho\)(_h_) distribution curve has an inflection point near 1.4 nm, which indicates a nonmonotonic decrease in the \(\rho\)(_h_) function in this area. This indicates a contribution of aggregate objects to the right wing of the distribution at \(h>\) 1.6 nm. Thus, the presence of HRP on the mica substrate surface is observed in the form of a mixture of monomers and aggregates.  
Figure 3C,D displays typical AFM images of the mica surface with HRP molecules, adsorbed from the solutions irradiated in the pulsed electromagnetic field for 40 min.  
As seen from Figure 3C,D, the appearance of objects with 3 to 4 nm height is observed on the substrate surface after its incubation in HRP solution, which was irradiated in PEMF for 40 min--in contrast to the case with the control solution. After processing the AFM data, a \(\rho\)(_h_) distribution was plotted analogously to the case with the control experiment. The resulting distribution plot is shown in Figure 3E (red curve). The obtained \(\rho\)(_h_) curve indicates that the maximum number of objects, visualized in the case of PEMF-irradiated HRP solution, have a height of about (2.8 \(\pm\) 0.2) nm, while the width at half-height makes up 3.5 nm. That is, after the irradiation of HRP enzyme solution in PEMF, the shape of the \(\rho\)(_h_) distribution curve changes significantly--in comparison with that obtained for the control HRP solution. The objects with heights from 2 to 4.5 nm, visualized on the surface of the substrate incubated in PEMF-irradiated solution, can be attributed to HRP aggregate structures. Thus, one can clearly observe an increase in the content of the aggregate structures adsorbed onto the mica surface from the HRP solution after its exposure to PEMF.