Biosensors and Bioelectronics 24 (2009) 2554–2558 Contents lists available at ScienceDirect Biosensors and Bioelectronics journal homepage: www.elsevier.com/locate/bios Fourier transform infrared immunosensors for model hapten molecules E. Gosselin a , M. Gorez a , M. Voué a,∗ , O. Denis b , J. Conti a , N. Popovic c , A. Van Cauwenberge c , E. Noel c , J. De Coninck a a Centre de Recherche en Modélisation Moléculaire, Université de Mons-Hainaut, Place du Parc, 20, B-7000 Mons, Belgium b Instituut Pasteur, WIV, Allergology Unit, Engelandstraat 642, B-1180 Brussels, Belgium c Hainaut Vigilance Sanitaire, Boulevard Sainctelette, 55, B-7000 Mons, Belgium article info Article history: Received 16 September 2008 Received in revised form 4 January 2009 Accepted 5 January 2009 Available online 14 January 2009 Keywords: Biosensors Ligand–receptor interaction FTIR Immunoreaction Monoclonal antibodies abstract We report experimental results concerning the detection of 2,4-dinitrophenol, under its free form or cou- pled to human serum albumin using Fourier transform infrared spectroscopy-based sensors. Competitive immunoreactions were carried out using several anti-dinitrophenol monoclonal antibodies. Comparison with enzyme-linked immunosorbent assays in competition is given for standard operating conditions. FTIR detection limits are comparable to those obtained by ELISA. The limits of detection are about 5–15ng/mL for the coupled DNP. Using the LO-DNP61 antibody, a detection limit of ∼ = 5 ng/mL was also estimated for the free DNP molecules but is much higher for the other antibodies. © 2009 Elsevier B.V. All rights reserved. 1. Introduction Sensors based on the molecular recognition of bio-molecules have already attracted intensive interest in many different fields such as medical diagnostics and control, environmental analysis, monitoring of biotechnological processes and quality control in food industry (see e.g. Andreescu and Sadik, 2004, and the refer- ences therein). In a recent article (Voué et al., 2007), we reviewed some specific aspects of biosensors and presented a new type of generic device suitable for the investigation of ligand–receptor interactions. These devices were based on the high sensitivity of Fourier transform infrared (FTIR) spectroscopy. This original method was based on the grafting of bifunctional spacer molecules directly at the surface of an internal reflection element (Fig. 1), made of silicon or germanium, avoiding the deposition of an intermedi- ate metal layer (Liley et al., 1997) or of an inorganic layer by sol–gel technique (Rigler et al., 2003; Onodera et al., 2007). Contrarily to surface plasmon resonance (SPR) or quartz crystal microbalance (QCM) sensors, FTIR sensors provide spectroscopic information concerning the chemical nature of the interacting molecules, as well as quantitative information concerning the amount of bound recep- tors and ligands. Furthermore, possible conformational transitions of the receptor during the interaction with its ligand can also be ∗ Corresponding author. Tel.: +32 65 373885/83; fax: +32 65 373881. E-mail address: michel.voue@crmm.umh.ac.be (M. Voué). monitored. This information is usually not accessible using stan- dard sensors, which only measure the mass loading of the surface. As the chemical structure of the interacting molecules is directly probed, FTIR-based sensors are de facto true label-free sensors. In this article, we apply the generic FTIR sensor technology to the making of FTIR-immunosensors. The principles of these sensors are the following: antigens coupled to proteins (typically, human or bovine serum albumin) are covalently immobilized at the sen- sor surface and the binding of mono- or poly-clonal antibodies is monitored as a function of time, after a pre-incubation step of the antibodies with some inhibitor molecules. These sensors can readily be compared to competition enzyme-linked immunosor- bent assays (ELISA). In particular, we have focused the research on the detection of small molecules: 2,4-dinitrophenol (DNP). Due to its low molecular weight, its chemical structure and its ability to be coupled with an immunogenic carrier, this molecule is often considered as an ideal model from hapten detection. The interest for such molecules is also due to their action as cellular metabolic poison with specific biological as well as pharmacological func- tions. 2,4-Dinitrophenol acts as a protein ionophore because of its ability to bind protons at one side of the plasma membrane and to release them at the other side. The consequence of this pro- ton transfer is that, at high concentrations of DNP in the cellular environment, it becomes impossible to maintain a proton gradi- ent with direct influence on the oxidative phosphorylation into mitochondrial membranes (see e.g. Sibille et al., 1995 and the ref- erences therein). For these reasons and its action on fatty acid synthesis (Rossmeisl et al., 2000), DNP has been used in the early 0956-5663/$ – see front matter © 2009 Elsevier B.V. All rights reserved. doi:10.1016/j.bios.2009.01.001