Gold-capped silicon for ultrasensitive SERS-biosensing: Towards human biofluids analysis.

Surface-enhanced Raman spectroscopy (SERS) has been widely used in a variety of biomedical, analytical, forensic and environmental investigations due to its chemical specificity, label-free nature combined with high sensitivity. Here, we report a simple method for the fabrication of reproducible and reliable, well-defined, stable SERS substrates with uniform and giant Raman enhancement suitable for routine trace chemical analysis and detection of biological compounds in complex biological fluids. We prepared porous silicone (PS) surface by a galvanostatic anodic etch of crystalline silicon wafers. The electrochemical process generates a specific layer of PS: the thickness and porosity of a given layer is controlled by the current density, the duration of the etch cycle, and the composition of the etchant solution. These substrates presented high sensitivity to p-mercaptobenzoic acid (p-MBA) at a low concentration of 10-6M and the enhancement factor of over 108 was achieved. Such high enhancement is attributed to semiconducting silicon-induced and stabilized hot spots. The uniform distribution of SERS-active 'hot-spots' on the Au/Si surface results in high reproducibility towards detecting p-MBA at 40 different, randomly selected positions on a single substrate (RSD=6.7%) and on twenty different SERS substrates prepared under identical conditions (RSD=8%). Designed substrates allow the ultrahigh sensitive and specific detection of human such biofluids as blood, urine and cerebrospinal fluid (CSF) in a reliable, label-free, and reproducible manner. The SERS spectra of these fluids are rich in patient-specific information and can be useful in many analytical and biomedical applications. We have shown that our developed SERS substrates allow the nanomolar detection of neopterin (bacterial infections' marker) in cerebrospinal fluid samples. In order to test the performance of our SERS method in term of low detection limit (LOD), the calibration curve i.e. plot of SERS intensity of the marker band at 695cm-1 versus the concentration of neopterin in CSF was constructed and used to calculate the neopterin concentration in clinical samples. The level of neopterin was significantly higher in CSF samples infected by Neisseria meningitidis, (54nmol/L), compared to normal (control) group, (4.3nmol/L). The high sensitivity, selectivity and stability of obtained SERS-active substrates combined with simple, low-cost, and easy method of producing offer a promising tool for SERS-based analysis in clinical trials.