WANG ET AL. VOL. XXX ’ NO. XX ’ 000–000 ’ XXXX www.acsnano.org A C XXXX American Chemical Society Enabling Rapid and Specific Surface-Enhanced Raman Scattering Immunoassay Using Nanoscaled Surface Shear Forces Yuling Wang, * ,†,§ Ramanathan Vaidyanathan, †,§ Muhammad J. A. Shiddiky, * ,† and Matt Trau * ,†,‡ † Centre for Personalized Nanomedicine, Australian Institute for Bioengineering and Nanotechnology (AIBN), Corner College and Cooper Roads (Bldg 75), and ‡ School of Chemistry and Molecular Biosciences, The University of Queensland, Brisbane QLD 4072, Australia. § Y.W. and R.V. contributed equally. P rotein immunoassays have widely been recognized as a powerful analy- tical tool for clinical diagnosis with fluorescence and enzyme-linked immuno- sorbent assay (ELISA)-based assays being widely used. Recently, surface-enhanced Raman scattering (SERS) immunoassays have gained widespread interest as an effective alternative to fluorescence and ELISA-based protein assays owing to their enhanced sensitivity, multiplex capability, and photostability. 13 However, the inte- gration of SERS immunoassays for clinical diagnosis is limited by two critical problems that include (i) longer incubation periods as a result of slow binding kinetics of heavier SERS particles to the target antigen (i.e., slow diffusion of particles) and (ii) high levels of nonspecific adsorption of molecules onto the sensor surface that reduce the specificity and sensitivity of detection. 49 Recently, Driskell et al. proposed an approach to increase the flux of antigen and SERS parti- cles to the solid-phase surface by using a rotated capture substrate. 10 The assay time was reduced from 24 h to 25 min in a 10-fold loss of sensitivity compared to the conven- tional SERS immunoassay. Furthermore, a syringe pump SERS immunoassay was developed to overcome diffusion-limited binding kinetics that often impedes rapid analysis in conventional SERS immuno- assay. 11 The assay time was reduced from 24 h to 10 min with a 10-fold improvement in detection limit. Despite these attempts being successful in reducing assay times, 10,11 nonspecific adsorption of nontarget mol- ecules still remains the biggest challenge in immunoassays, 12,13 in particular, SERS-based immunoassays, 3,9 and very few attempts have been reported to address this long- standing problem. To circumvent this pro- blem, herein, we propose an innovative platform that utilizes nanoscaled alternat- ing current electrohydrodynamic (ac-EHD)- induced surface shear forces to enhance the capture efficiency (i.e., increase diffusion of target and SERS particles) as well as * Address correspondence to y.wang27@uq.edu.au, m.shiddiky@uq.edu.au, m.trau@uq.edu.au. Received for review March 31, 2015 and accepted May 15, 2015. Published online 10.1021/acsnano.5b01929 ABSTRACT A rapid and simple approach is presented to address two critical issues of surface-enhanced Raman scattering (SERS)-based immunoassay such as removal/avoiding nonspecific adsorption and reducing assay time. The approach demonstrated involves rationally designed fluorophore-integrated gold/ silver nanoshells as SERS nanotags and utilizes alternative current electrohy- drodynamic (ac-EHD)-induced nanoscaled surface shear forces to enhance the capture kinetics. The assay performance was validated in comparison with hydrodynamic flow and conventional immunoassay-based devices. These nanos- caled physical forces acting within nanometer distances from the electrode surface enabled rapid (40 min), sensitive (10 fg/mL), and highly specific detection of human epidermal growth factor receptor 2 in breast cancer patient samples. We believe this approach presents potential for the development of rapid and sensitive SERS immunoassays for routine clinical diagnosis. KEYWORDS: SERS . immunoassay . ac-EHD . shear force . nonspecific adsorption ARTICLE