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1 November 2002 Molecular probes for two-photon excited fluorescence and second harmonic generation imaging of biological membranes
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Novel microscopies based on nonlinear optical (NLO) phenomena are attracting increasing interest in the biology community owing to their potentialities in the area of real-time, non-damaging imaging of biological systems. In particular, second-harmonic generation (SHG) and two-photon excited fluorescence (TPEF) are NLO phenomena that scale with excitation intensity squared, and thus give rise to an intrinsic 3-dimensional resolution when used in microscopic imaging. In this perspective, we have implemented a molecular engineering approach toward NLO-probes specifically designed for SHG and/or TPEF imaging of cellular membranes. We have designed nanoscale rod-like fluorophores showing very large TPEF cross-sections in the visible red, outperforming standard fluorophores such as fluorescein by up to two orders of magnitude. Bolaamphiphilic derivatives combining high TPEF cross-sections and affinity for cellular membranes were prepared. Their incorporation into model or cell membranes can be monitored by TPEF microscopy. Amphiphilic push-pull chromophores showing both high TPA and SHG cross-sections in the near-IR region were designed as NLO-probes for imaging of biological membranes by simultaneous SHG and TPEF microscopy. These NLO-phores offer intriguing potentialities for imaging of fundamental biological processes such as adhesion, fusion or for reporting of membrane electrical potentials.
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Laurent Porres, Olivier Mongin, Bharath K. G. Bhatthula, Mireille H. Blanchard-Desce, Lionel Ventelon, Laurent Moreaux, T. Pons, and Jerome Mertz "Molecular probes for two-photon excited fluorescence and second harmonic generation imaging of biological membranes", Proc. SPIE 4812, Nonlinear Spectroscopy, (1 November 2002);

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