Surface plasmon resonance combined with spectroscopic ellipsometry read-out for probing surface–biomolecule interaction Maria M. Giangregorio ⁎, Giuseppe V. Bianco, Pio Capezzuto, Giovanni Bruno, Maria Losurdo Institute of Inorganic Methodologies and of Plasmas, CNR-IMIP, University of Bari, via Orabona 4, 70126 Bari, Italy abstract article info Available online xxxx Keywords: Ellipsometry Surface plasmon resonance Gold nanoparticles Hemin Thiol Albumin Spectroscopic ellipsometry combined with localized surface plasmon resonance (LSPR) of gold nanoparticles (Au NPs) is exploited to design label-free bionsensors. We demonstrate that the size of Au NPs significantly affects the sensitivity of the ellipsometry analysis. Additionally, functionalizing Au NPs of different sizes with molecules/proteins of different sizes and shapes, such as dodecanethiol, hemin, human albumin and its antibody, we show that the size of nanoparticles can strongly influence the binding activity of adsorbed proteins and, consequently, the sensor functioning. Specifically, Au NPs with a diameter in the range 30–50 nm exhibit higher sensitivity to the change in the optical properties, and the variations of the ellipsometric parameter Ψ allow discerning phenomena of aggregation of Au NPs of the sensor, of detachment of Au NPs and of protein chemisorp- tion on Au NPs. The data are discussed in terms of two main factors affecting the ellipsometry sensitivity, i.e., the dependence of the LSPR electromagnetic enhancement on the Au NP size, and the strength of the interaction of the functionalizing molecule with Au NPs. © 2013 Elsevier B.V. All rights reserved. 1. Introduction Localized surface plasmon resonance (LSPR) sensors are one of the most investigated optical sensing tools used for label-free detection of various chemical and biological molecules [1]. In this context, total internal reflection ellipsometry (TIRE) exploiting the LSPR effect of a thin gold film has demonstrated a high sensitivity of the Δ ellipsometric parameter in measuring the interaction between bio-molecules also in opaque media [2,3]. Indeed, since Natan's group in 1998 showed that gold nanoparticles (Au NPs) yield over 25 times higher sensitivity than a conventional LSPR device [4], there has been an increasing interest in exploiting Au NP incorporation into SPR devices to amplify the signal and improve sensitivity. Au NPs have a high surface-to- volume ratio that can provide a higher density of active sites for binding events [5]; they also generate enhanced electromagnetic fields that affect the local environment and give the nanoparticles an enhanced reactivity. Therefore, Au NP LSPR sensors have the potential of enhanc- ing the detection of immobilized biomolecules in the picoMolar range [6]. An additional advantage of NP LSPR compared to propagating evanescent SPR in thin metallic films is that the NP size and shape allow tuning the LSPR wavelength, amplitude and the field decay length, l d . The decay length of NP enhanced field is approximately 5–15 nm or 1–3% of the light wavelength and differs greatly from the 200–300 nm decay length (approximately 15–25% of the light wavelength) of a SPR sensor [7]. The shorter LSPR electromagnetic field decay length could provide enhanced surface sensitivity of the LSPR sensor by minimizing bulk medium contributions [8,9]. The elec- tromagnetic field decay lengths for NPs increase with the increase of the NP size [10]. As an example, sensors fabricated from Au NPs with a 39 nm diameter exhibited maximum sensitivity to the change of local refractive index by biotin–streptavidin [11]. For the quantification of the “analytical volume” also the dimension of the analyte with respect to the Au NPs size is important. If the decay length is small with respect to the analyte/biomolecule thickness, the response to the binding event could be weak. If, on the other hand, the decay length is large compared to the dielectric layer thickness, the analyte will occupy a small fraction of the sensing volume, resulting again in a weak response. Thus, for an optimal response, a transducer has to be designed considering that the Au NP size and LSPR field enhancement decay length maximize the re- sponse to a given analyte–receptor system [11,12]. Additionally, in the case of protein adsorption onto nanoparticles, it should also be consid- ered that, when the size of nanoparticles becomes comparable to the di- mensions of the proteins, there may be significant effects of local steric hindrance on binding mechanisms and functionality of the adsorbed protein. A recent study has shown that the structure/stability/function of the adsorbed protein can be a surprisingly strong function of the size of the nanoparticle onto which it is adsorbed [13]. This motivates further the investigation of the degree of stabilization/destabilization of mole- cules/proteins depending on both size and shape of nanoparticles and size, shape, orientation, and nature of proteins of organic molecules. In this paper, we analyze the different sensitivities of the Ψ and Δ ellipsometric parameters to the interaction of Au NPs of different sizes with molecules of different sizes, shapes/structures and reactivities, i.e., the hemin, the dodecanethiol and the human albumin (HSA) and its antibody IgC. Thin Solid Films xxx (2013) xxx–xxx ⁎ Corresponding author. E-mail address: michelaria.giangregorio@ba.imip.cnr.it (M.M. Giangregorio). TSF-32993; No of Pages 6 0040-6090/$ – see front matter © 2013 Elsevier B.V. All rights reserved. http://dx.doi.org/10.1016/j.tsf.2013.11.143 Contents lists available at ScienceDirect Thin Solid Films journal homepage: www.elsevier.com/locate/tsf Please cite this article as: M.M. Giangregorio, et al., Thin Solid Films (2013), http://dx.doi.org/10.1016/j.tsf.2013.11.143