A SERS-Active Electrospun Polymer Mesh for Spatially Localized pH Measurements of the Cellular Microenvironment

Abstract

Extracellular pH (pHe) is an important chemical factor in many cellular processes and disease pathologies. The routine sampling of pHe in vitro could lead to innovative advances in therapeutics. To this end, we have fabricated a novel gold-coated polymer mesh, which facilitates the real-time measurement of pHe via surface-enhanced Raman scattering (SERS). In this proof of concept study, we apply our SERS sensor to measure metabolically induced changes in the pHe of carcinoma-derived cell line HepG2/C3A. We demonstrate that gold-coated polyurethane electrospun nanofibers (AuNF) have strong and reproducible SERS spectra of surface-adsorbed analytes. By functionalizing AuNF with pH-responsive reporter 4-mercaptobenzoic acid (MBA), we have developed an accurate pH SERS sensor for the extracellular microenvironment. We cultured HepG2/C3A on the surface of MBA-AuNF and measured an acidic shift in pHe at the cell−fiber interface. Following exposure to staurosporine, an apoptosis-inducing drug, we observed changes in the HepG2/C3A cellular morphology indicative of controlled cell death, and detected an increase in the pHe of HepG2/C3A. These results demonstrate how subtle changes in pHe, induced by the metabolic activity of cells, can be measured with our novel SERS sensor MBA-AuNF. The excellent pH measurement performance of MBA-AuNF provides a unique platform to study extracellular pH on the microscale and will help to deepen our understanding of pHe in disease pathology.

SERS microsensors for pH measurements in the lumen and ECM of stem cell derived human airway organoids

“…We functionalised the gold surface, via thiol-gold bonding, with 4mercaptobenzoic acid (MBA), a pH-sensitive reporter molecule previously applied to SERS pH sensing in the cellular environment. 3,15,16 SEM images of SERS-MS showed a monolayer of gold nanoparticles formed on the microparticle surface and reflected bright field microscopy confirmed the presence of surface adsorbed AuNPs on SERS-MS in solution (Fig. 1 and Fig.…”

“…We functionalised the gold surface, via thiol-gold bonding, with 4-mercaptobenzoic acid (MBA), a pH-sensitive reporter molecule previously applied to SERS pH sensing in the cellular environment. 3,15,16…”

“…Previous studies have demonstrated MBA-based SERS pH sensing is influenced by the chemistry of the local ECM environment. 15,17 By incubating SERS-MS in the ECM for 14 days we simulate the time the sensors spent in the ECM during AO maturation and ensured the accuracy of in situ SERS-MS pH measurements in the ECM and lumen of live AOs. MBA-based SERS pH sensing employs the intensity of the pH-sensitive v s (COO) peak at ∼1430 cm −1 relative to the aromatic ring deformation peak ( v (ref)) at ∼1590 cm −1 to measure pH in the aqueous environment surrounding the sensor (Fig.…”

Optical Sensing Strategies for Probing Single-Cell Secretion

“…SERS has a sensitivity comparable to that of fluorescence, but is less influenced by photobleaching. A simple yet efficient approach for SERS analysis of H + excretion from living cells was developed by directly culturing cells on the SERS substrates premodified with pH-sensitive Raman probes such as 4-mercaptopyridine and 4-mercaptobenzoic acid. − The detection was based on the deprotonation and protonation of the probe molecules at different pH conditions, which resulted in a detectable change of their SERS spectra. − To avoid the effect of ion diffusion and dilution in the extracellular space, the strategy of tethering probes on the cell surface was also feasible for SERS analysis. For example, a cell membrane-anchored SERS biosensor was designed for measuring extracellular pH at micrometer-scale resolution .…”

Floating Ag-NPs@Cu-NW bundles fabricated on copper mesh for highly sensitive SERS detection of uric acid in pretreatment-free urine

“…Recently, commercial metal fiber materials such as CM have been used as skeleton structures to construct SERS substrates due to their cost-effectiveness, large surface area, and flexibility. [18][19][20][21][22] When using CM as a substrate, Cu(OH) 2 nanowires can be grown on its surface by different methods. On one hand, Cu(OH) 2 nanowire arrays are nanostructures capable of facile and high-throughput synthesis with a controllable morphology, and can be fabricated in large areas on a copper substrate.…”