Biosensors and Bioelectronics 20 (2005) 1539–1548 Development of a fluid functionalized lipidic matrix applied to direct in situ polynucleotide detection Daphn´ e L. Thomas, Lo¨ ıc J. Blum, Agn` es P. Girard-Egrot ∗ LaboratoiredeG´ enieEnzymatiqueetBiomol´ eculaire,UMR5013/EMB2—CNRS/UCBL,Universit´ eClaudeBernardLyon1,43Bddu11novembre1918, F-69622VilleurbanneCedex,France Received 28 May 2004; received in revised form 15 July 2004; accepted 15 July 2004 Available online 13 September 2004 Abstract This work presents a new approach for direct detection of polyelectrolytes at the air–water interface, based on the investigation of the interfacial properties of an active lipidic matrix especially designed for polynucleotide immobilization. A synthetic lipid with a cationic spermine headgroup, DiOctadecylamidoGlycylSpermine (DOGS), was spread at the interface to form a distortable film able to capture polynucleotides. The control of the organization state of this functionalized monolayer upon compression was achieved by recording surface pressure–area (π–A) isotherm diagrams, presenting a specific shape with a typical liquid expanded–liquid condensed phase transition on a pure water subphase. In the presence of various dsDNA concentrations in the subphase, the isotherms were markedly modified in a time and concentration-dependent manner. The main modifications, corresponding to a large shift towards higher molecular areas and a clear fading of the phase transition, were corroborated by the fine analysis of the monolayer compressibility profile, thus suggesting a characteristic change in the monolayer fluidity as a function of both time and DNA concentration. Moreover, an ATR-Fourier transform infrared (ATR-FTIR) characterization showed evidences for the adsorption of DNA strands onto the lipidic matrix. The direct observation of the mixed monolayer morphology by Brewster angle microscopy (BAM) strongly suggests that DNA adsorption induces a reorganization of lipids at the interface, as evidenced by the change in the condensed lipidic domains morphology in the presence of DNA in the subphase. The immobilization of various polynucleotidic probes of 4000, 400 and 22 base length, confirmed by fluorescence microscopy, had similar effects on DOGS interfacial properties. Preliminary studies are finally presented to explore the possibility of using this system for the study of hybridization between complementary strands. Hence, this study demonstrates this functionalized matrix behaves as a fluid support where polynucleotide immobilization induces interfacial properties modifications, which could be further exploited through the experimental characterization of Faraday instabilities sensitive to visco-elasticity variations. © 2004 Elsevier B.V. All rights reserved. Keywords: DNA detection; Functionalized interface; Polynucleotide immobilization; Langmuir monolayer 1. Introduction In the wide field of DNA biosensors technology, going from the development of new sensing devices to elaborated DNA chips, the development of DNA-modified surfaces lies in the heart of numerous research studies. As demonstrated with the great variety of solid materials, selected either for their role as a support bearing biomolecules including syn- ∗ Corresponding author. Tel.: +33 472 448 532; fax: +33 472 447 970. E-mailaddress: agnes.egrot@univ-lyon1.fr (A.P. Girard-Egrot). thetic polymers, glass (Fodor et al., 1991; Beier and Hoheisel, 1999; Yershov et al., 1996), or their active role in the detec- tion step, such as silicon (Lamture et al., 1994; Blanchard et al., 1996; Strother et al., 2000; Patole et al., 2003; Chrisey et al., 1996) and metal surfaces (Xu and Bard, 1995; Frutos et al., 1998), tremendous researches are being performed to op- timize the immobilization of probes on a substrate, or finding alternative methods for detecting hybridized probes. Another aspect of nanobiotechnology concerns the elaboration of self-assembled nanoscale systems, where biomolecules are used as elementary structures to form a 0956-5663/$ – see front matter © 2004 Elsevier B.V. All rights reserved. doi:10.1016/j.bios.2004.07.027