SERS-based Immunoassay in a Microfluidic System for the Multiplexed Recognition of Interleukins from Blood Plasma: Towards Picogram Detection

SERS-active nanostructures incorporated into a microfluidic device have been developed for rapid and multiplex monitoring of selected Type 1 cytokine (interleukins: IL-6, IL-8, IL-18) levels in blood plasma. Multiple analyses have been performed by using nanoparticles, each coated with different Raman reporter molecules: 5,5'-dithio-bis(2-nitro-benzoic acid) (DTNB), fuchsin (FC), and p-mercatpobenzoic acid (p-MBA) and with specific antibodies. The multivariate statistical method, principal component analysis (PCA), was applied for segregation of three different antigen-antibody complexes encoded by three Raman reporters (FC, p-MBA, and DTNB) during simultaneous multiplexed detection approach. To the best of our knowledge, we have also presented, for the first time, a possibility for multiplexed quantification of three interleukins: IL-6, IL-8, and IL-18 in blood plasma samples using SERS technique. Our method improves the detection limit in comparison to standard ELISA methods. The low detection limits were estimated to be 2.3 pg.ml^-1, 6.5 pg.ml^-1, and 4.2 pg.ml^-1 in a parallel approach, and 3.8 pg.ml^-1, 7.5 pg.ml^-1, and 5.2 pg.ml^-1 in a simultaneous multiplexed method for IL-6, IL-8, and IL-18, respectively. This demonstrated the sensitivity and reproducibility desirable for analytical examinations.