Improvement of Surface-Enhanced Raman Scattering Method for Single Bacterial Cell Analysis

Abstract

Surface-enhanced Raman scattering (SERS) is a useful tool for label-free analysis of bacteria at the single cell level. However, low reproducibility limits the use of SERS. In this study, for the sake of sensitive and reproducible Raman spectra, we optimized the methods for preparing silver nanoparticles (AgNPs) and depositing AgNPs onto a cell surface. We found that fast dropwise addition of AgNO 3 into the reductant produced smaller and more stable AgNPs, with an average diameter of 45 ± 4 nm. Compared with that observed after simply mixing the bacterial cells with AgNPs, the SERS signal was significantly improved after centrifugation. To optimize the SERS enhancement method, the centrifugal force, method for preparing AgNPs, concentration of AgNPs, ionic strength of the solution used to suspend the cells, and density of the cells were chosen as impact factors and optimized through orthogonal experiments. Finally, the improved method could generate sensitive and reproducible SERS spectra from single Escherichia coli cells, and the SERS signals primarily arose from the cell envelope. We further verified that this optimal method was feasible for the detection of low to 25% incorporation of 13 C isotopes by the cells and the discrimination of different bacterial species. Our work provides an improved method for generating sensitive and reproducible SERS spectra.

Unmasking Bacterial Identities: Exploiting Silver Nanoparticle ‘Masks’ for Enhanced Raman Scattering in the Rapid Discrimination of Diverse Bacterial Species and Antibiotic-Resistant Strains

“…Traditional nanoparticles, for example, face challenges in achieving complete and uniform adsorption on bacterial surfaces. Instead, they can only adhere to selected regions of the bacterial surface, leading to the acquisition of partial SERS information. ,− This limitation is particularly evident with centrifuged nanoparticles or nanoparticles modified with a capping agent, which enhance in situ bacterial signals but yield similar Raman signals across different bacterial species. Consequently, bacteria can only be identified and differentiated by statistical analysis as amplified Raman signals exhibit homogeneous properties driven primarily by protein or nucleic acid signals. , This homogeneity obscures the specific identity of bacterial strains, complicating efforts to accurately determine the species and resistance profiles.…”

Simple and Inexpensive Fabrication of High Surface-Area Paper-Based Gold Electrodes for Electrochemical and Surface-Enhanced Raman Scattering Sensing

“…Alternatives have emerged to overcome this drawback, with surface-enhanced Raman scattering (SERS) being a popular approach that can result in signal enhancements of up to 10 14 compared to classic Raman spectroscopy. 18 Materials with surfaces rich in pores, edges, and kinks can enhance the electromagnetic field of the incident radiation and result in a significant SERS effect. 19 Research to develop materials for SERS has focused on gold and silver nanostructures, since these possess the most significant enhancement factor.…”

“…Yet, the Raman scattering efficiency is low, with only 1 out of 10 8 photons typically scattered by the sample, which translates into low sensitivity. Alternatives have emerged to overcome this drawback, with surface-enhanced Raman scattering (SERS) being a popular approach that can result in signal enhancements of up to 10 14 compared to classic Raman spectroscopy . Materials with surfaces rich in pores, edges, and kinks can enhance the electromagnetic field of the incident radiation and result in a significant SERS effect .…”