In this chapter, various techniques are reviewed with focus on the identification of complex biological agents such as bacteria, viruses, proteins, and enzymes using SERS-active silver substrates. Biological targets have multiple peculiarities that add to the challenges of the SERS biosensing. In regards to the direct non-labeled sensing of bacteria, it was discovered that all bands in the registered SER spectra were generated by metabolites released from bacterial cells. It undermined the prior notion of non-labeled detection and identification of bacteria based on the presumed spectra of cellular walls. However, it also provides new opportunities for the SERS analysis of bacteria. The SERS measurements of viruses can be performed with SERS-active surfaces or colloidal solutions of silver nanoparticles. However, the use of surfaces requires extensive sample preparation and often lacks sensitivity, while colloidal SERS substrates have another problem—most types of silver nanoparticles are negatively charged and have a poor interaction with likewise predominantly negatively charged virions. Thus, a challenge is posed to develop SERS-ready positively charged silver nanoparticles or use other methods to enforce the non-specific binding of viruses to the silver surfaces. Meanwhile, SER spectra of proteins are nearly impossible to acquire at adequate sensitivity. Thus, non-direct measurements are the only way. SERS provides the most benefits when working with relatively small molecules, so small molecules serving as Raman probes can be used as an intermediary to produce SER spectra. For enzymes like butyrylcholinesterase, it means measuring SER spectra of substrates and products of the relevant reaction, while for other proteins, specialized techniques must be developed. It can be concluded that biological targets require a case-by-case approach. Prior experiences with direct SERS measurements of highly Raman-active molecules like R6G and others often used in fundamental studies might not be relevant in bioanalytics.
SERS for Bacteria, Viruses, and Protein Biosensing