Optical scattering artifacts observed in the development of multiplexed surface enhanced Raman spectroscopy nanotag immunoassays.

Here we describe scattering based signal suppression artifacts encountered while developing multiplex lateral flow (LF) immunoassay using surface enhanced Raman spectroscopy (SERS) "nanotags" as analyte labels. Using these SERS nanotags, we have produced a quantitative test for inflammation biomarkers that is transferable to the point of care (POC). The SERS assay shows similar performance when compared with a fluorescent nanoparticle POC test. Here, using cardiac and inflammation biomarkers, we highlight the need to carefully optimize the concentration of assay components when using SERS nanotags and a single-line multiplexing approach. We show that in certain circumstances the SERS signal may be suppressed, leading to a significant underestimation of the analyte concentrations. Using electron microscopy and optical spectroscopy, we demonstrate that the error in the measurement is associated with the light scattering properties of the nanotags. These findings will be applicable to other nanoparticle labels with high light scattering coefficients. Through careful modification of the assay to reduce the impact of light scattering, it is possible to produce quantitative assays, but potentially at the expense of multiplexing capability and assay sensitivity.