ИСТИНА

SUMMARY The effectiveness of a multimodal sensor lies in its ability to utilize the complementary advantages of different detec-tion methods. This study demonstrates this by combining Fabry-Perot interference and surface-enhanced Raman spectroscopy (SERS) on a unified substrate—gold and silver-decorated porous silicon nanowires (AuAg@pSi NWs). The total reflection spectra of AuAg@pSi NWs were characterized by the presence of interference fringes occurring in the thin films upon reflection of white light at the upper and lower boundaries of the nanowire arrays. After bacterial adsorption, the distance between the interference peaks in the spectra changes due to the change in the samples ef-fective optical thickness (EOT). At the same time, monitoring of bacteria adsorbed on AuAg@pSiNWs was accom-plished using SERS. The resulting spectrum exhibited distinct scattering bands attributed to glycosidic bond vibrations within the peptidoglycan of the bacterial cell wall, as well as valence vibrations of the C-N bond within the adenine por-tion of the lipid layer constituents of the cell wall. Thus, the developed sensor demonstrates potential for rapid label-free diagnosis and identification of bacteria, establishing it as a versatile tool for diverse applications in the field of mi-crobial detection. 1. INTRODUCTION Optical biosensors have significant advantages over other analytical methods due to good sensitivity, convenience and ease of use, reproducibility and reliability. To convert the signal in optical sensors, one can use the effects of interfer-ometry, surface plasmon resonance, diffraction grating of photonic crystals, converters based on optical waveguides, ellipsometry, etc. [1]. The most common optical sensors are based on the effects of light interference in thin silicon nanostructure films such as porous silicon of various morphologies [2] or silicon nanowires (SiNWs) [3]. The principle of operation of such a sensor is that the illumination of thin silicon nanostructure films with white light leads to reflection of light at the top and bottom interfaces of films, creating Fabry–Perot interference, where the interference frequency is determined by the EOT of the silicon nanostructure films [2,4]. The change in the effective refractive index of the silicon nanostructure films after the adsorption of biological molecules and cells manifests itself in a shift of the interference fringes and/or a change in their amplitude [2–7]. Another popular method for detecting various biomolecules is SERS [8]. A simple method for decorating of SiNWs with gold and silver particles by reducing them from AgNO3 and AuCl3 in the presence of 5M HF in order to impart SERS-active properties to the resulting composite substrates was presented [9]. Furthermore, bilirubin was successfully de-tected with a detection limit of 1 µM using SERS on Au-decorated SiNWs [10], as well as internalin B, a protein asso-ciated with the pathogenic bacteria Listeria monocytogenes, was successfully detected using SERS on Ag-decorated SiNWs [11]. In this study two methods of detection, Fabry–Perot interference and SERS, were combined and it was shown that SiNWs covered with Au and Ag nanoparticles can be used for creation of bimodal optical sensor for bacteria diagnos-tics. 2. EXPERIMENTAL RESULTS AND DISCUSSIONS PSi NWs coated with metal particles were obtained by MACE from p-type c-Si wafer with crystallographic orientation (100) and resistivity of 1–5 mΩ·cm. To decorate the resulting pSi NWs with noble metal nanoparticles, the etched wa-fer was placed alternately in solutions with AgNO3 and AuCl3. Figure 1 a,b shows SEM micrographs of obtained AuAg@pSi NWs (top and side views). The top view shows that the outer surface of the silicon nanostructures is evenly covered with silver and gold nanoparticles (shown as light dots in the image). From the side view, one can judge the ordering of SiNW arrays with a predominant orientation along the (100) crystallographic direction. The SiNW layer thickness is 2.3 μm, and the thickness of the bimetallic layer is 100 nm. Figure 1 c,d shows SEM micrographs (top and side views) of L. Innocua adsorbed on AuAg@pSi NWs at a con-centration of 3.2·107 CFU/mL. It has been established that bacteria are located both on the surface of SiNWs and in the space between nanowires, which contributes to the achievement of a better optical signal when they are detected by measuring total reflection and SERS. The total reflectance spectra of AuAg@pSi NWs were characterized by the presence of interference fringes as a result of Fabry-Perot interference in thin films upon reflection of white light at the top and bottom interfaces of AuAg@pSi NWs. Interference fringes is related to the EOT of AuAg@pSi NWs by equation (1): EOT = 2Lneff = mλ, (1) where m is the spectral order, λ is the wavelength of light, L is the thickness of AuAg@pSi NWs and neff is the effective refractive index of AuAg@pSi NWs (nanowire diameter and distance between the nanowires are less than the light wavelength). After the adsorption of bacteria, the distance between the interference peaks changes due to the change in neff and, accordingly, the EOT. SERS spectra of L. Innocua gives bacterial concentration dependence within the range from 0.6·107 to 6.4·107 CFU/mL from which it can be seen that bacterial identification is appropriate. The most intense Raman bands in the acquired spectrum correspond to cell wall components of gram-positive bacteria Listeria. Thus, a technique has been developed for obtaining composite AuAg@pSi NWs for fabrication of a bimodal optical sensor based on them for diagnosing bacteria using L. Innocua as an example. Scheme of operation of the sensor element is shown in figure 2. Two different modalities are shown. The interference occurs due to reflections reflection of white light at the top and bottom interfaces of AuAg@pSi NWs. SERS becomes possible due to the decoration of silicon nanowires with noble metals and the penetration of bacteria into the structure. Fig. 1. SEM micrographs. a) Top view of obtained AuAg@pSi NWs; b) side view of obtained AuAg@pSi NWs; c) top view of L. Innocua adsorbed on AuAg@pSi NWs at a concentration of 3.2·107 CFU/mL; d) side view of L. Innocua adsorbed on AuAg@pSi NWs at a concentration of 3.2·107 CFU/mL. Fig.2. Scheme of operation of a bimodal sensor based on AuAg@pSi NWs. 3. CONCLUSIONS AuAg@pSi NWs with a SiNWs thickness of 2.3 μm and a bimetallic layer thickness of 100 nm were obtained and stud-ied. The possibility of diagnosing L. Innocua up to concentrations of 0.6·107 CFU/mL by changes of EOT of AuAg@pSi NWs has been demonstrated. The possibility of diagnosing L. Innocua by SERS spectra after their adsorption on AuAg@pSi NWs up to concentrations of 0.6 107 CFU/mL has been demonstrated. Scattering bands are observed in the SERS spectra, which are characteristic of proteins in the cell wall of gram-positive Listeria bacteria. It can be con-cluded that AuAg@pSi NWs are effective as a bimodal optical sensor for diagnosing bacteria. The developed sensor demonstrates potential for rapid label-free diagnosis and identification of bacteria, establishing it as a versatile tool for diverse applications in the field of microbial detection. ACKNOWLEDGMENT The study was supported by the Russian Science Foundation, grant No. 22-72-10062 https://rscf.ru/en/project/22-72-10062/ and was performed using SEM of the Training Methodical Center of Lithography and Microscopy, Moscow State University. REFERENCES [1] M. Nirschl, F. Reuter, J. Vörös, Biosensors, 2011, 1(3), 70. [2] A. Jane, R. Dronov, A. Hodges, N. H. Voelcker. Trends in biotech., 2009, 27(4), 230. [3] K. A. Gonchar, S. N. Agafilushkina, D. V. Moiseev, I. V. Bozhev, A. A. Manykin, E. A. Kropotkina, A. S. Gambaryan, L. A. Osminkina, Mater. Res. Express, 2020, 7, 035002. [4] V. S. Lin, K. Motesharei, K. P. Dancil, M. J. Sailor, M. R. Ghadiri, Science, 1997, 278(5339), 840. [5] M. B. Gongalsky, A. A. Koval, S. N. Schevchenko, K. P. Tamarov, L. A. Osminkina. J. Electr. Soc., 2017, 164(12), B581. [6] N. Massad-Ivanir, G. Shtenberg, E. Segal, J. Vis. Exp., 2013, 81. [7] N. Massad-Ivanir, G. Shtenberg, N. Raz, C. Gazenbeek, D. Budding, M.P. Bos, E. Segal, Sci. 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