We detected concentration-dependent surface-enhanced Raman scattering (SERS) spectra of several label-free proteins (lysozyme, ribonuclease B, avidin, catalase, and hemoglobin) for the first time in aqueous solutions. Acidified sulfate was used as an aggregation agent to induce high electromagnetic enhancement in SERS. Strong SERS spectra of simple and conjugated protein samples could easily be accessed after the pretreatment with the aggregation agent. The detection limits of the proposed method for lysozyme and catalase were as low as 5 microg/mL and 50 ng/mL, respectively. This detection protocol for label-free proteins has combined simplicity, sensitivity, and reproducibility and allows routine qualitative and relatively quantitative detections. Thus, it has great potential in practical high-throughput protein detections.

In this paper, a direct and simple detection method based on surface-enhanced Raman scattering (SERS) named "heat-induced SERS sensing method" is proposed for rapid determination of glutathione in aqueous solutions. It was found that highly enhanced SERS spectra of glutathione can be obtained if the silver colloids adsorbed with the analyte were heated up before the SERS measurement. Besides, it was revealed that silver particles with a size of approximately 60 nm are suitable for this study and that the SERS intensity is also influenced by the dropped sample volume, drying temperature, buffer concentration, and pH of the solution. It is noted that the thiol group of glutathione has a particularly strong interaction with a silver surface compared with other small biological molecules without a thiol group, validating this method to detect glutathione selectively. Under the optimal conditions, the detection of glutathione can be finished within 5 min, and the detection limit of ca. 50 nM can be reached, which is much better than the reported detection limit of glutathione (approximately 1 microM) by SERS. The enhancement factor of the proposed heat-induced SERS sensing method for the detection of glutathione is about 7.5 x 10(6). The proposed method holds a specific selectivity toward glutathione, facilitating its rapid detection in practical applications.

Surface-enhanced Raman scattering (SERS) enhancement and the reproducibility of the SERS signal strongly reflect the quality and nature of the SERS substrates because of diverse localized surface plasmon resonance (LSPR) excitations excited at interstitials or sharp edges. LSPR excitations are the most important ingredients for achieving huge enhancements in the SERS process. In this report, we introduce several gold and silver nanoparticle-based SERS-active substrates developed solely by us and use these substrates to investigate the influence of LSPR excitations on SERS. SERS-active gold substrates were fabricated by immobilizing colloidal gold nanoparticles on glass slides without using any surfactants or electrolytes, whereas most of the SERS-active substrates that use colloidal gold/silver nanoparticles are not free of surfactant. Isolated aggregates, chain-like elongated aggregates and two-dimensional (2D) nanostructures were found to consist mostly of monolayers rather than agglomerations. With reference to correlated LSPR and SERS, combined experiments were carried out on a single platform at the same spatial position. The isolated aggregates mostly show a broadened and shifted SPR peak, whereas a weak blue-shifted peak is observed near 430 nm in addition to broadened peaks centered at 635 and 720 nm in the red spectral region in the chain-like elongated aggregates. In the case of 2D nanostructures, several SPR peaks are observed in diverse frequency regions. The characteristics of LSPR and SERS for the same gold nanoaggregates lead to a good correlation between SPR and SERS images. The elongated gold nanostructures show a higher enhancement of the Raman signal than the the isolated and 2D samples. In the case of SERS-active silver substrates for protein detection, a new approach has been adopted, in contrast to the conventional fabrication method. Colloidal silver nanoparticles are immobilized on the protein functionalized glass slides, and further SERS measurements are carried out based on LSPR excitations. A new strategy for the detection of biomolecules, particularly glutathione, under aqueous conditions is proposed. Finally, supramolecular J-aggregates of ionic dyes incorporated with silver colloidal aggregates are characterized by SERS measurements and correlated to finite-difference time-domain analysis with reference to LSPR excitations.