## 3 Results and Discussion
### Evaluation of the Mobility of DNA-GNP Associates by Agarose Gel Electrophoresis
These circumstances should facilitate the complementary interaction of adsorbed FL molecules with R molecules adsorbed on another GNP. As a result, highly charged but large and \"slow\" associates (FL/GNP + R/GNP) can come into being. With a further increase in the concentration of FL and R at 45 \({}^{\circ}\)C, the same trends were observed as those at 37 \({}^{\circ}\)C: the proportion of \"fast\" associates increased from 36% to 74% (Figure 7B, for 45 \({}^{\circ}\)C, row \"% bottom\"). The presence of large and \"slow\" associates (i.e., a redistribution of some oligonucleotides from filled GNPs to naked GNPs) was confirmed in experiments involving (i) separation of mixture (FL + R) /GNPs (Figure 7B) and of a mixture of associates obtained beforehand (FL-GNP + R-GNP) (Figure S5A) and (ii) mixing of additional citrated GNPs with ssDNA/GNP associates, followed by incubation for 0, 2, and 24 h (Figure S5A,B). Analysis by means of radiolabeled T20 clearly confirmed the redistribution of some oligonucleotides from loaded GNPs to naked GNPs (Figure 8). The initial ratio of the amount of T20 adsorbed on the particles of the upper and lower fractions (according to electrophoretic mobility) was 21% and 79% (Figure 8A,B, lane T20-GNP). Immediately after the addition of the naked GNPs to T20*/GNPs, this ratio shifted to 29% and 71%, respectively (Figure 8A,B, lane T20/GNP+GNP, 0 h). After 24 h, the amounts of DNA in the upper and lower particle fractions equal  
Figure 8: Agarose gel electrophoresis of T20/GNP associates and their mixtures with naked GNPs immediately after the mixing (0 h) and after incubation for 24 h. A gel scan (**A**) and a gel autoradiograph (**B**). Naked GNPs and initial T20/GNP associates were applied to the gel as controls for the mobility and distribution of the associates. 50% each (Figure 8A,B, lane T20/GNP+GNP, 24 h). Moreover, immediately after mixing, we noticed desorption of some T20 molecules (22% or 7 out of 32 molecules), and after 24 h, almost no desorption was detectable (6% or 2 out of 32 molecules). In other words, upon addition of naked GNPs, there was probably desorption of some DNA molecules from the surface of the initial associates into the solution. These molecules were then adsorbed onto the surface of the added GNPs. As a consequence, after 24 h, DNA desorption into the solution was not detectable. Therefore, after the mixing of two types of associates (FL/GNP and R/GNP), desorption of the appreciable proportion of DNA into the solution at 1 day after the mixing indicates the formation of a duplex by these DNAs in solution (Figure S6). Thus, the ratios of the capacity and of electrophoretic mobility--of GNP associates with ssDNA or with a mixture of complementary ssDNA--indirectly confirm possible formation of DNA duplexes upon adsorption on GNPs at all tested concentrations and temperatures. It is known that the method of DNA binding to the surface of GNPs affects the size of the associate and its mobility in an agarose gel [50]. Indirectly, the existence of such a relation has been demonstrated for associates of GNPs with thiol-containing and native oligonucleotides. The associates of GNPs with thiol-containing ssDNA have a larger radius as compared to the associates of GNPs with thiol-dsDNA or with thiol-free ssDNA [49]. As a consequence, associates of GNPs with thiol-bound dsDNA migrate more slowly in an agarose gel than do associates of GNPs with noncovalently bound dsDNA [50].