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Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections

Theodossiou, TA; Thrasivoulou, C; Ekwobi, C; Becker, DL; (2006) Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections. BIOPHYS J , 91 (12) 4665 - 4677. 10.1529/biophysj.106.093740. Green and gold open access

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Abstract

We performed second harmonic generation (SHG) imaging of collagen in rat-tendon cryosections, using femtosecond laser scanning confocal microscopy, both in backscattering and transmission geometries. SHG transmission images of collagen fibers were spatially resolved due to a coherent, directional SHG component. This effect was enhanced with the use of an index-matching fluid (n(l) 1.52). The average SHG intensity oscillated with wavelength in the backscattered geometry (isotropic SHG component), whereas the spectral profile was consistent with quasi-phase-matching conditions in transmission geometry (forward propagating, coherent SHG component) around 440 nm (lambda(p) 880 nm). Collagen type I from bovine Achilles tendon was imaged for SHG in the backscattered geometry and its first-order effective nonlinear coefficient was determined (vertical bar d(eff)vertical bar approximate to 0.085(+/- 0.025) x 10-(12)mV(-1)) by comparison to samples of inorganic materials with known effective nonlinear coefficients (LiNbO3 and LiIO3). The SHG spectral response of collagen type I from bovine Achilles tendon matched that of the rat-tendon cryosections in backscattered geometry. Collagen types I, II, and VI powders (nonfibrous) did not show any detectable SHG, indicating a lack of noncentrosymmetric crystalline structure at the molecular level. The various stages of collagen thermal denaturation were investigated in rat-tendon cryosections using SHG and bright-field imaging. Thermal denaturation resulted in the gradual destruction of the SHG signal.

Type:Article
Title:Second harmonic generation confocal microscopy of collagen type I from rat tendon cryosections
Open access status:An open access publication. A version is also available from UCL Discovery.
DOI:10.1529/biophysj.106.093740
Publisher version:http://www.ncbi.nlm.nih.gov/pmc/ articles/PMC1779934/?tool=pubmed
Keywords:OPTICAL HARMONIC-GENERATION, DIFFERENTIAL SCANNING CALORIMETRY, THERMAL-DENATURATION, TAIL TENDON, 2-PHOTON-EXCITED FLUORESCENCE, BIOLOGICAL TISSUES, REFRACTIVE INDICES, VISUAL OBSERVATION, LASER-PULSES, POLARIZATION
UCL classification:UCL > School of Life and Medical Sciences > Faculty of Life Sciences > Biosciences (Division of) > Cell and Developmental Biology

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