Browsing by Subject "Second harmonic microscopy"

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    Enhancement of second harmonic microscopy images in collagen-based thick samples using radially polarized laser beams
    (Elsevier, 2021-07-07) Martínez Ojeda, Rosa María; Hernández García, Carlos; Bueno García, Juan Manuel; Física
    The visualization of collagen-based tissues imaged with second harmonic generation (SHG) microscopy strongly depends on the polarization state of the scanning laser beam. This has been often explored using spatially homogeneous states (mainly linear or circular). However, vector beams (i.e. laser beams with spatially non-uniform polarization states) have become a powerful tool in different research field and technological applications. In particular, radially polarized vector beams are known to provide a tight focal spot and lateral resolution enhancement. Here we propose the use of radially polarized beams to improve the quality of SHG microscopy images. The performance of this particular type of vector beam is compared to that obtained with a circularly polarized light beam for different non-stained specimens and depth locations. Results show that independently of the sample’s thickness, the efficiency of SHG processes is increased when using the radially polarized beam, also leading to a higher image quality. Our results open the route for using vector beams to get a better visualization of biological details within the imaged areas, especially at deeper locations.
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    Quantitative structural organization of the sclera in chicks after deprivation myopia measured with second harmonic generation microscopy
    (Frontiers Media, 2024-10-22) Bueno García, Juan Manuel; Martínez-Ojeda, Rosa M.; Fernández, Enrique J.; Feldkaemper, Marita; Física
    Visual deprivation causes enhanced eye growth and the development of myopia, which is associated with a change in the arrangement of collagen fibers within the sclera. A second harmonic generation (SHG) microscope has been used to image the collagen fibers of unstained scleral punches from the posterior part of chicken eyes. We aimed to analyze the fibrous scleral tissue and quantify the changes in collagen organization in relation to the extent of induced deprivation myopia. The scleral architecture was assessed with the Radon transform (RT) through the parameter called structural dispersion (SD) that provides an objective tool to quantify the level of organization of the collagen network. We found that final refraction and axial length changes were linearly correlated. However, no significant differences in scleral thickness were found for different amounts of induced myopia. In contrast, a significant correlation between SD and refraction was demonstrated, ranging from a non-organized (in the control sclerae) to a quasi-aligned distribution (with a dominant direction of the fibers, in the sclera of myopic chicks). These findings demonstrate a remodeling process of the scleral collagen associated with myopia progression that can be measured accurately combining SHG imaging microscopy and RT algorithms.