| Description |
Surface-enhanced Raman scattering (SERS) is a technique that can be used for the detection of materials down to the single molecule level. The development of the extrinsic Raman Label (ERL) that incorporates biorecognition, a Raman reporter molecule, and gold nanoparticles (AuNPs), is the base for an extensible, sensitive, and selective SERS-based sandwich immunoassay. The use of SERS as a quantitative detection platform, however, has not progressed past the laboratory. Research presented here in describes how the analysis and production of a SERS immunoassay substrate has a significant role in the reliability and reproducibility of SERS substrates. First, the analysis of the SERS immunoassay platform and can be simulated as a random distribution of points on a surface. Simulation results indicated the best method to improve the accuracy of the analysis was through increasing the number of measurements or increasing the area measured by a single measurement. The precision of the measurement, however, was only improved by increasing the analysis area. This indicates that a larger laser spot used for analysis improves the accuracy and precision of SERS measurements. Second, to produce a SERS substrate with a random distribution of ERLs, the adsorption of ERLs should follow diffusional transport to increase the uniformity of ERLs on the substrate. By inverting the substrates during the ERL incubation step, sedimentation of the ERLs is directed away from the substrate and stable ERLs left in suspension diffuse to the substrate. Diffusional transport and a more even distribution of ERLs increased the reliability and reproducibility of the SERS substrates. Improved SERS immunoassay techniques implemented in conjunction with a novel pretreatment of serum samples for tuberculosis (TB) diagnostics was used to validate the SERS method. The TB marker mannose-capped lipoarabinomannan (ManLAM) is a lipopolysaccharide cell wall component that is constantly sloughed off the surface of the virulent bacterium, Mycobacterium tuberculosis. Normally ManLAM complexes with serum protein inhibiting detection but the use of a simple five step pretreatment method frees ManLAM improving the limit of detection (LoD). Improved SERS methodologies and sample pretreatment provide promising sensitivity and specificity for set of patient samples. |
