Nanoscale Chemical Imaging of Human Cell Membranes Using Tip‐Enhanced Raman Spectroscopy

Abstract

Lack of appropriate tools for visualizing cell membrane molecules at the nanoscale in a non‐invasive and label‐free fashion limits our understanding of many vital cellular processes. Here, we use tip‐enhanced Raman spectroscopy (TERS) to visualize the molecular distribution in pancreatic cancer cell (BxPC‐3) membranes in ambient conditions without labelling, with a spatial resolution down to ca. 2.5 nm. TERS imaging reveals segregation of phenylalanine‐, histidine‐, phosphatidylcholine‐, protein‐, and cholesterol‐rich BxPC‐3 cell membrane domains at the nm length‐scale. TERS imaging also showed a cell membrane region where cholesterol is mixed with protein. Interestingly, the higher resolution TERS imaging revealed that the molecular domains observed on the BxPC‐3 cell membrane are not chemically “pure” but also contain other biomolecules. These results demonstrate the potential of TERS for non‐destructive and label‐free imaging of cell membranes with nanoscale resolution.

Regioselective Tip-Enhanced Raman Spectroscopy of Lipid Membranes with Sub-Nanometer Axial Resolution

“…Therefore, like the DPPC bilayer, we assign the prominent 2908 and 2966 cm −1 bands observed in the TERS spectra of the cell surface to the t+ and c + vibrations of the CH 3 groups in the choline moiety. In our previous work, 48 we detected the ν C−N vibrational mode of the choline moiety in the TERS measurements of the DPPC bilayers and pancreatic cancer cells, which further supports our hypothesis that TERS spectra of the lipid membranes measured in this work represent vibrations in the choline moiety of phospholipids.…”

“…To check the reproducibility of these results, we performed TERS measurements with an additional TERS probe at five more locations in a different part of the DPPC bilayer as shown in Figure S4a,b, where TERS spectra with the exact same shape were observed. The confocal Raman spectra of the same region of the DPPC bilayer were measured by lifting the TERS probe 200 μm away from the sample surface . However, no Raman signal was detected in the confocal Raman spectra, as shown in Figure S4c, confirming that the TERS signals of the DPPC bilayer originated from the near field.…”

Nanoscale chemical analysis of 2D molecular materials using tip-enhanced Raman spectroscopy

“…Furthermore, the instrument/sample drift is a significant issue in hyperspectral TERS imaging, as highlighted in recent works. 32,57 Therefore, the step size of TERS imaging should be selected carefully depending on the size of the region of interest. For TERS measurements of homogeneous samples such as uniform supported lipid monolayers or bilayers, a large step size (up to 50 nm) can be an effective method to obtain an overview of the sample.…”

“…16 In the last two decades, tip-enhanced Raman spectroscopy (TERS) has emerged as a powerful analytical technique that combines the high spatial resolution of AFM or STM and the chemical sensitivity and specificity of surface-enhanced Raman spectroscopy (SERS) to study surface chemistry at the nanoscale in a label-free and non-destructive manner. [16][17][18][19] TERS has been successfully applied for nanoscale analysis in several areas of scientific research such as 2D materials including graphene, MoS 2 and WSe 2 and graphitic sheets, [20][21][22][23][24] onsurface decomposition chemistry, 25 polymer blends, 26 photocatalytic reactions, [27][28][29] organic solar cells, 30 supported lipid membranes, 31 biomembranes, 32 and biological cells. 33 In TERS, Raman scattering from a nanoscopic volume of the sample is plasmonically enhanced by several orders of magnitude owing to a highly intense and localised electromagnetic (EM) field, which is generated at the apex of a metallic SPM probe via a combination of localised surface plasmon resonance and lightening rod effect.…”

Toward the Next Frontiers of Vibrational Bioimaging

“…The collective oscillation of electrons induced in LSPR confines the incident light field into a nanoscale near-field around AFM probes with up to 10 8 signal enhancements and leads to mode-specific vibrational imaging with atomic resolution . For bioimaging, TERS has been used to collect in situ Raman signatures from biological species, including amyloid fibrils in water and mammalian cell membranes . Combining the SERS or TERS with coherent Raman excitation would in principle lead to even higher sensitivity.…”