## 3 Results and Discussion  
The presence of F atoms as doping agent in ZnO powders strongly affected all the intrinsic properties of ZnO nanocrystals, in particular its morphology, by inducing the formation of a clear granular nanometric phase [16]. This effect was well evident when the fluorinated nZnO-F and the undoped nZnO films were directly grown on solid substrates and studied for comparison. Both nZnO-F and nZnO films were grown on flat crystalline silicon by a hydrothermal synthesis, which required a seed layer of ZnO sputtered on the substrates before the synthesis. Due to the nanometric features of these samples, their morphological characterization was performed by SEM imaging. The results, reported in Figure 1, highlighted an impressive difference of the morphology in the different materials: The nZnO-F appeared as a packed film constituted by flake-like nanograins of about 600 nm; while the nZnO was formed by standard hexagonal nanocolumns of 200 nm in diameter almost perpendicular to the plane. The surface roughness of doped and undoped nZnO was measured by a profilometric technique; the nZnO-F was characterized by a rms (root mean squared) of \(20\pm 2\) nm, while the nZnO surface result was smoother (rms of only \(9\pm 1\) nm). The two samples were functionalized with FITC-labeled PrA (PrA*) following the chemical procedure described in Scheme 1. PrA was used as a model molecule since it has great relevance from a sensing point of view and for the biosensors surface chemical modification. PrA is often covalently coupled to the surface of transducer materials by several methods which span from direct adsorption to click chemistry [17; 18; 19]. PrA can be found in nature as a virulence factor specific for Staphylococcus aureus, bound to glycans on the bacterial cell surface. PrA is also able to link to the von Willebrand factor that is a protein for the hemostasis process. Moreover, more important is the fact that PrA selectively binds the Fc-region of human antibodies, thus resulting essentially in the correct orientation of the antibodies fixed on a support surface. The chemical modification strategy of nZnO and nZnO-F was based on two steps. In the first one, the covalent binding of a FITC-labelled PrA was monitored by the FT-IR spectroscopy, WCA analysis, and fluorescence imaging in order to assess the silanization of a zinc oxide nanostructured surface. The hydrothermal grown zinc oxide had a hydrophilic surface after the synthesis, which was made in aqueous solution (see Scheme 1 (1)). The presence of OH groups promoted the Zn-O-Si bounds after the silanization with APTES, resulting in an amino-terminated surface (Scheme 1 (2)). The BS\({}^{3}\) crosslinker and the exposure to a PrA solution completed the functionalization procedure  
Figure 1: Representative SEM images of nanostructured fluorine (F)-doped ZnO (nZnO-F) (**A**) and nZnO (**B**). (Scheme 1 (3) and (4)). In the second step, increasing concentrations of PrA were grafted on both surfaces in search of which was better functionalized by the same procedure. Figure 2 shows the FT-IR spectra of the nZnO-F acquired before and after the last functionalization step with PrA*; spectra related to intermediate functionalization steps (i.e., APTES and BS3) were not reported because any substantial variation was not observed. The features at 1540 and 1400 cm\({}^{-1}\) were attributed to asymmetric and symmetric stretching modes of the acetate groups (-COO) absorbed during the synthesis process [20]. The peaks at 1650 cm\({}^{-1}\) and about 3300 cm\({}^{-1}\), only visible in the spectrum of the functionalized sample, were due to the amide I band and to N-H stretching vibration of protein, respectively [21]. Differently, the FTIR spectrum of nZnO, after the PrA* immobilization procedure, did not present any evident change, which corresponded to a very low amount of protein bound on its surface (data not shown here).