## 3 Results and Discussion
### Evaluation of Ribosomes and Polysomes Adsorption on Functionalized Gold Surfaces
#### 3.2.2 Polysome Adsorption  
Similarly to ribosomes (Section 3.2.1), polysome adsorption was analyzed on gold and gold-functionalized surfaces. Fraction 10 of purified polysomes (Figure S1) was selected for this study. Fraction 10 is a medium-heavy weighted fraction, comprising polysomes formed by 6-10 ribosomes each [25]. Examples of AFM images of these polysomes are shown in Figure 1, while the morphological distribution resulting from grain-analysis is reported in Figure S7 (data and fit), and in Table 4 (summary of fitted peaks). In addition, in the case of polysomes, the morphological characteristics measured by AFM were compared to reference conditions [25]. After deposition on gold, polysomes were imaged as globular or rod-shaped isolated clusters of ribosomes (about ten ribosomes per cluster) occasionally connected by strand-like features, which likely correspond to naked mRNA (Figure 1, panel A). The distributions of morphological parameters appear to be segregated in two main populations: a first population represented by smaller objects with an height of 5.9 nm, width of 24.9 nm and length of 33.4 nm, and a second population that comprises larger objects with an height of 12.6 nm, width of 87.2 nm and length of 122.5 nm. These values show a quite good conservation in polysome shape and distributions compared with the reference. Similarly, polysomes on MCOOH functionalized gold retained similar size values in the population of bigger objects (height of 13.6 nm, width of 76.6 nm and length of 107.6 nm). On MNH\({}_{2}\) surfaces was also noticed the presence of small strand-like structures. Moreover, the distributions obtained from depositing on MNH\({}_{2}\) surfaces, required three Gaussian components for a meaningful fit, evidencing an highly abundant population of smaller objects, as defined by the three studied properties. Finally, polysomes deposited on neutral surfaces (i.e., MOH and M11 CH\({}_{3}\)) were found to be severely disrupted, particularly on the hydrophobic M11 CH\({}_{3}\) surface (Figure S8). The favorable adhesion of ribosomes and polysomes we found on charged surfaces (both positively and negatively charged) can be elicited by the presence on the ribosomal surface of several charged groups, as shown in the Figure S3. Moreover, we note that during the adhesion phase, relatively high concentration of divalent cations are present in polysome preserving buffers, which can also favor the indirect adhesion of negatively charged groups on negatively charged surfaces [31; 32]. \begin{table}
\begin{tabular}{c c c c c c c c c} \hline \hline \multicolumn{1}{c}{**Surface**} & \multicolumn{5}{c}{**Ribosome Density (obj. \(\times\) 10\({}^{8}\)/cm\({}^{2}\))**} & **Polysome Density (obj. \(\times\) 10\({}^{8}\)/cm\({}^{2}\))** \\ \hline bare gold & \(117\pm 13\) & \(15\pm 4\) & \\ \hline MCOOH & \(84\pm 4\) & \(19\pm 5\) & \\ \hline MNH\({}_{2}\) & \(132\pm 11\) & \(30\pm 6\) & \\ \hline MOH & \(52\pm 12\) & \(14\pm 7\) & \\ \hline M11 CH\({}_{3}\) & n.d. & n.d. & \\ \hline \hline \end{tabular}
\end{table} 
Table 3: Density of ribosomes and polysomes adsorbed on the different surfaces. \begin{table}
\begin{tabular}{c c c c c c c c c c} \hline \hline \multicolumn{1}{c}{**Surface**} & \multicolumn{5}{c}{**(nm, Center/Width)**} \\ \hline
**Surface** & **H1** & **H2** & **H3** & **W1** & **W2** & **W3** & **L1** & **L2** & **L3** \\ \hline bare gold & - & 5.9/1.9 & 12.6/1.7 & - & 24.9/7.5 & 87.2/18.0 & - & 33.4/9.7 & 122.5/19.2 \\ \hline MCOOH & - & 8.3/2.1 & 13.6/0.6 & - & 29.5/3.9 & 76.6/14.1 & - & 35.6/3.7 & 107.6/28.9 \\ \hline MNH\({}_{2}\) & 4.0/0.8 & 7.5/2.3 & 14.4/1.6 & 14.4/5.7 & 27.6/9.2 & 80.7/21.5 & 19.0/4.6 & 34.7/11.0 & 110.8/32.1 \\ \hline \hline \end{tabular}
\end{table} 
Table 4: Height (H1, H2, H3) and lateral size values (width—W1, W2, W3; length—L1, L2, L3) obtained form the Gaussian fits of the morphological value distributions of polysomes deposited on bare and functionalized gold (center and width of Gaussian fits are shown). Data and fits are reported in Figure S7.