Surface enhanced Raman scattering of living cells can be used to study molecule–nanostructure interactions and biochemical processes in vivo .

The ultraviolet resonance Raman (UVRR) spectra of the two proteins bovine serum albumin (BSA) and human serum albumin (HSA) in an aqueous solution are compared with the aim to distinguish between them based on their very similar amino acid composition and structure and to obtain signals from tryptophan that has only very few residues. Comparison of the protein spectra with solutions of tryptophan, tyrosine, and phenylalanine in comparative ratios as in the two proteins shows that at an excitation wavelength of 220 nm, the spectra are dominated by the strong resonant contribution from these three amino acids. While the strong enhancement of two and one single tryptophan residue in BSA and HSA, respectively, results in pronounced bands assigned to fundamental vibrations of tryptophan, its weaker overtones and combination bands do not play a major role in the spectral range above 1800 cm ^–1 . There, the protein spectra clearly reveal the signals of overtones and combination bands of phenylalanine and tyrosine. Assignments of spectral features in the range of Raman shifts from 3800 to 5100 cm ^–1 to combinations comprising fundamentals and overtones of tyrosine were supported by spectra of amino acid mixtures that contain deuterated tyrosine. The information in the high-frequency region of the UVRR spectra could provide information that is complementary to near-infrared absorption spectroscopy of the proteins.

Gold nanostars are a versatile plasmonic nanomaterial with many applications in bioanalysis. Their interactions with animal cells of three different cell lines are studied here at the molecular and ultrastructural level at an early stage of endolysosomal processing. Using the gold nanostars themselves as substrate for surface-enhanced Raman scattering, their protein corona and the molecules in the endolysosomal environment were characterized. Localization, morphology, and size of the nanostar aggregates in the endolysosomal compartment of the cells were probed by cryo soft-X-ray nanotomography. The processing of the nanostars by macrophages of cell line J774 differed greatly from that in the fibroblast cell line 3T3 and in the epithelial cell line HCT-116, and the structure and composition of the biomolecular corona was found to resemble that of spherical gold nanoparticles in the same cells. Data obtained with gold nanostars of varied morphology indicate that the biomolecular interactions at the surface in vivo are influenced by the spike length, with increased interaction with hydrophobic groups of proteins and lipids for longer spike lengths, and independent of the cell line. The results will support optimized nanostar synthesis and delivery for sensing, imaging, and theranostics.

Infrared nano‐spectroscopy (nano‐IR) using scattering‐type near‐field optical microscopy (s‐SNOM) is becoming an important tool for analyzing vibrational spectra in nanometer‐scale volumes of intact samples. Recent s‐SNOM experiments using ultrathin membranes have enabled nanoscopy of cells and biomolecules in liquid environments. This study reports the use of SiC membranes as a stable, biocompatible interface between cells in their medium and the atomic force microscopy (AFM) probe of the s‐SNOM, ensuring broadband nano‐IR experiments above 1050 cm ⁻¹ . Nano‐IR images and spectra are collected from fibroblast cells grown adherently to confluence on the membranes using a broadband synchrotron IR source, probing the membrane‐adjacent cellular region. Optimized tip‐sample interaction allows to significantly increase the signal‐to‐noise‐ratio in nano‐IR imaging. The alteration of the tapping amplitude and set‐point of the AFM cantilever allows to adjust the probing depth and suggests a nano‐IR tomography approach that uses a single demodulation harmonic of the signal. The nano‐IR spectra of fibroblast cells resemble their far‐field spectra, but reflect a heterogeneity in the composition and structure of proteins, nucleic acids, carbohydrates, and of membrane lipid organization. The results demonstrate nano‐IR probing of complex samples in liquid media and suggest ways to improve efficiency and standardization of existing approaches in vibrational nanoscopy.