## 3 Results and Discussion
### Silicon-Based Surfaces for Polysomes Capture  
Results obtained with gold surfaces were then used to set-up a purification route based on the use of silicon surfaces, which are more suitable for the fabrication of microdevices. In particular, based on the final observations in Section 3.2.2, functionalization protocols inducing negatively charged surfaces were tested. TG-SO surfaces were treated with two different organic silanes, CST and triCAS (Table S1), containing 1 and 3 carboxyl functional residues, respectively. The functionalized surfaces were characterized via AFM and XPS before monitoring the polysomes adsorption.  
Figure 1: 2 x 2 \(\upmu\)m\({}^{2}\) AFM images of medium-heavy weighted polysome fractions deposited on bare gold (**A**) and MCOOH (**B**), MNH\({}_{2}\) (**C**) and MOH (**D**) functionalized gold. The z-range color-scale is 15 nm.  
The AFM analysis showed that initial TG-SO surfaces are quite smooth, with an average roughness \(S_{a}\) of (\(0.18\pm 0.01\)) nm. This roughness does not increase significantly after functionalization with either triCAS or CST. Moreover, XPS analysis confirmed that the silanization process occurred successfully, since all the silanized surfaces showed a larger amount of carbon content. In particular, 2 to 3 times more carbon was measured on the silanized surfaces compared to bare TG-SO (Table S2). These results demonstrate that the functionalization process resulted in a coverage of the pristine surface.  
The unmodified (but plasma cleaned) and the modified surfaces were tested for their capacity of adsorption of polysomes, as in the above Section 3.2.2. Medium-high heavy polysomes were incubated on plasma-treated TG-SO and on the functionalized TG-SO surfaces. AFM imaging (Figure 2) showed a similar result for polysomes adsorbed on plasma-treated TG-SO (Figure 2, panel A) and on surfaces treated with triCAS (Figure 2, panel B), i.e., a fair number of adsorbed polysomes without any marked shape of degradation. Concerning polysomes entrapped on CST- treated TG-SO (Figure 2, panel C), a certain degree of degradation can be noticed, as also confirmed by the analysis of morphological distributions. The three morphological properties indeed showed a decrease in the peak center, compared to the TG-SO and triCAS surfaces (Table 5 and Figure S9). Furthermore, about two thirds of the detected objects are estimated to contribute to the population of smaller objects, characterized by low width and length values (Figure S9).  
The overall distribution of fitting peaks obtained using triCAS (Table 5) quite closely resembles the results obtained with MCOOH-treated gold (see Table 4).  
However, on surfaces functionalized with triCAS, some isolated smaller objects are present, as reflected by the appearance of a peak at 4.5 nm in the height distribution, which is instead absent in the MCOOH-gold surfaces. Summarizing, by comparing the density of objects adsorbed on surfaces (Table 6), the TG-SO silanized with triCAS is the surface able to retain more objects, yielding a result comparable to that found employing MCOOH-functionalized gold surfaces. TG-SO functionalized with CST yields comparable results in term of density, but a high fraction of these structures are possibly degraded polysomes. Therefore, silicon surface functionalized with triCAS are the negatively-charged surface most promising for polysome capture.  
Figure 2: 2 \(\times\) 2 μm2 AFM images of polysomes adsorbed on negatively charged silicon-based surfaces. Panel **A**: plasma-treated TG-SO; panel **B**: triCAS treated silicon surface; panel **C**: CST treated TG-SO (Section 2.2.2).