24 September 2009 Measurement of the quadratic hyperpolarizability of the collagen triple helix and application to second harmonic imaging of natural and biomimetic collagenous tissues
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Abstract
Collagen is a major protein of the extracellular matrix that is characterized by triple helical domains. It plays a central role in the formation of fibrillar and microfibrillar networks, basement membranes, as well as other structures of the connective tissue. Remarkably, fibrillar collagen exhibits efficient Second Harmonic Generation (SHG) so that SHG microscopy proved to be a sensitive tool to probe the three-dimensional architecture of fibrillar collagen and to assess the progression of fibrotic pathologies. We obtained sensitive and reproducible measurements of the fibrosis extent, but we needed quantitative data at the molecular level to further process SHG images. We therefore performed Hyper- Rayleigh Scattering (HRS) experiments and measured a second order hyperpolarisability of 1.25 10-27 esu for rat-tail type I collagen. This value is surprisingly large considering that collagen presents no strong harmonophore in its aminoacid sequence. In order to get insight into the physical origin of this nonlinear process, we performed HRS measurements after denaturation of the collagen triple helix and for a collagen-like short model peptide [(Pro-Pro- Gly)10]3. It showed that the collagen large nonlinear response originates in the tight alignment of a large number of weakly efficient harmonophores, presumably the peptide bonds, resulting in a coherent amplification of the nonlinear signal along the triple helix. To illustrate this mechanism, we successfully recorded SHG images in collagenous biomimetic matrices.
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A. Deniset-Besseau, A. Deniset-Besseau, M. Strupler, M. Strupler, J. Duboisset, J. Duboisset, P. De Sa Peixoto, P. De Sa Peixoto, E. Benichou, E. Benichou, C. Fligny, C. Fligny, P.-L. Tharaux, P.-L. Tharaux, G. Mosser, G. Mosser, P.-F. Brevet, P.-F. Brevet, M.-C. Schanne-Klein, M.-C. Schanne-Klein, "Measurement of the quadratic hyperpolarizability of the collagen triple helix and application to second harmonic imaging of natural and biomimetic collagenous tissues", Proc. SPIE 7487, Optical Materials in Defence Systems Technology VI, 74870E (24 September 2009); doi: 10.1117/12.829882; https://doi.org/10.1117/12.829882
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