Encapsulation of lysozyme within graphene sandwich reduces its thermal denaturation during SERS-analysis

We report on a novel approach for the encapsulation of vital antibacterial protein lysozyme between two graphene-containing films on a solid substrate composed of silver nanoparticles on porous silicon (Ag/PSi) possessing surface-enhanced Raman scattering (SERS) activity. SERS- spectra of 10 nM lysozyme were collected and compared for the graphene-free Ag/PSi, the graphene-coated Ag/PSi, and the graphene sandwich with lysozyme on Ag/PSi. The spectral regions with characteristic bands of the protein were subjected to quantitative/qualitative analysis. The findings revealed that the bottom graphene layer facilitated a more ordered orientation of protein molecules, augmented the Raman signal due to graphene-induced charge transfer, and dissipated heat generated by plasmonic nanoparticles/laser. The top graphene layer additionally reduced the temperature effect on the protein by several degrees, which was verified by computer modeling. Thus, it was demonstrated that the graphene sandwich enabled recording of SERS-spectra of the protein with the slightly changed secondary structure. Moreover, the graphene coating resulted in more protein Raman bands compared to that of the uncoated Ag/PSi. Therefore, the encapsulation of lysozyme within the graphene sandwich on a plasmonic substrate reduces its thermal carbonization, enabling the reliable analysis of proteins by SERS- spectroscopy at concentrations that are clinically relevant.