• Title/Summary/Keyword: Optical Microscopy

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A Quantitative Study of the Quality of Deconvolved Wide-field Microscopy Images as Function of Empirical Three-dimensional Point Spread Functions

  • Adur, Javier;Vicente, Nathalie;Diaz-Zamboni, Javier;Izaguirre, Maria Fernanda;Casco, Victor Hugo
    • Journal of the Optical Society of Korea
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    • v.15 no.3
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    • pp.252-263
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    • 2011
  • In this work, for the first time, the quality of restoration in wide-field microscopy images after deconvolution was analyzed as a function of different Point Spread Functions using one deconvolution method, on a specimen of known size and on a biological specimen. The empirical Point Spread Function determination can significantly depend on the numerical aperture, refractive index of the embedding medium, refractive index of the immersion oil and cover slip thickness. The influence of all of these factors is shown in the same article and using the same microscope. We have found that the best deconvolution results are obtained when the empirical PSF utilized is obtained under the same conditions as the specimen. We also demonstrated that it is very important to quantitatively check the process' outcome using several quality indicators: Full-Width at Half-Maximum, Contrast-to-Noise Ratio, Signal-to-Noise Ratio and a Tenengrad-based function. We detected a significant improvement when using an indicator to measure the focus of the whole stack. Therefore, to qualitatively determinate the best deconvolved image between different conditions, one approach that we are pursuing is to use Tenengrad-based function indicators in images obtained using a wide-field microscope.

Visualization of Epidermis and Dermal Cells in ex vivo Human Skin Using the Confocal and Two-photon Microscopy

  • Choi, Sang-Hoon;Kim, Wi-Han;Lee, Yong-Joong;Lee, Ho;Lee, Weon-Ju;Yang, Jung-Dug;Shim, Jong-Won;Kim, Jin-Woong
    • Journal of the Optical Society of Korea
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    • v.15 no.1
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    • pp.61-67
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    • 2011
  • The confocal laser scanning microscopy and two-photon microscopy was implemented based on a single laser source and an objective lens. We imaged and compared the morphology of identical sites of ex vivo human skin using both microscopes. The back-scattering emission from the sample provided the contrast for the confocal microscopy. The intrinsic autofluorescence and the second harmonic generation were used as the luminescence source for the two-photon microscopy. The wavelength of the Ti:Sapphire laser was tuned at 710 nm, which corresponds to the excitation peak of NADH and FAD in skin tissue. The various cell layers in the epidermis and the papillary dermis were clearly distinguished by both imaging modalities. The two-photon microscopy more clearly visualized the intercellular region and the nucleus of the cell compared to the confocal microscopy. The fibrous structures in the dermis were more clearly resolved by the confocal microscopy. Numerous cells in papillary dermal layer, as deep as $100\;{\mu}m$, were observed in both CLSM and two-photon microscopy. While most previous studies focused on fibrous structure imaging (collagen and elastin fiber) in the dermis, we demonstrated that the combined imaging with the CLSM and two-photon microscopy can be applied for the non-invasive study of the population, distribution and metabolism of papillary dermal cells in skin.

Comparison of Existing Methods to Identify the Number of Graphene Layers

  • Sharbidre, Rakesh Sadanand;Lee, Chang Jun;Hong, Seong-Gu;Ryu, Jae-Kyung;Kim, Taik Nam
    • Korean Journal of Materials Research
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    • v.26 no.12
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    • pp.704-708
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    • 2016
  • The unique characteristics of graphene make it an optimal material for crucial studies; likewise, its potential applications are numerous. Graphene's characteristics change with the number of total layers, and thus the rapid and accurate estimation of the number of graphene layers is essential. In this work, we review the methods till date used to identify the number of layers but they incorporate certain drawbacks and limitations. To overcome the limitations, a combination of these methods will provide a direct approach to identify the number of layers. Here we correlate the data obtained from Raman spectroscopy, optical microscopy images, and atomic force microscopy to identify the number of graphene layers. Among these methods, correlation of optical microscopy images with Raman spectroscopy data is proposed as a more direct approach to reliably determine the number of graphene layers.

Identification and Correction of Microlens-array Error in an Integral-imaging-microscopy System

  • Imtiaz, Shariar Md;Kwon, Ki-Chul;Alam, Md. Shahinur;Hossain, Md. Biddut;Changsup, Nam;Kim, Nam
    • Current Optics and Photonics
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    • v.5 no.5
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    • pp.524-531
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    • 2021
  • In an integral-imaging microscopy (IIM) system, a microlens array (MLA) is the primary optical element; however, surface errors impede the resolution of a raw image's details. Calibration is a major concern with regard to incorrect projection of the light rays. A ray-tracing-based calibration method for an IIM camera is proposed, to address four errors: MLA decentering, rotational, translational, and subimage-scaling errors. All of these parameters are evaluated using the reference image obtained from the ray-traced white image. The areas and center points of the microlens are estimated using an "8-connected" and a "center-of-gravity" method respectively. The proposed approach significantly improves the rectified-image quality and nonlinear image brightness for an IIM system. Numerical and optical experiments on multiple real objects demonstrate the robustness and effectiveness of our proposed method, which achieves on average a 35% improvement in brightness for an IIM raw image.

Lateral Resolution Enhancement in Confocal Self-interference Microscopy with Commercial Calcite Plate

  • Kang DongKyun;Yoo HongKi;Lee SeungWoo;Gweon Dae-Gab
    • Journal of the Optical Society of Korea
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    • v.9 no.1
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    • pp.32-35
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    • 2005
  • In light microscopy, spatial resolution is limited by diffraction effect. Confocal microscopy has improved resolutions in both lateral and axial directions, but these are still limited by diffraction effect. Confocal self-interference microscopy (CSIM) uses interference between two perpendicularly polarized beams to enhance lateral resolution. In previous research, we proposed a calcite plate with its optic-axis perpendicular to the propagation angle and one of the boundary surfaces of the plate. This type of plate is not widely used to our knowledge. In this paper, we change the calcite plate to more common one, which is commercially available. This calcite plate has its optic axis in the plane of incidence. We analyze the characteristics of this calcite plate and numerically compare the performances of CSIM in previous research and CSIM with the commercial calcite plate. Numerical results show improved performance when using the commercial calcite plate

Polarization State of Scattered Light in Apertureless Reflection-mode Scanning Near-Field Optical Microscopy

  • Cai, Yongfu;Aoyagi, Mitsuharu;Emoto, Akira;Shioda, Tatsutoshi;Ishibashi, Takayuki
    • Journal of Magnetics
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    • v.18 no.3
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    • pp.317-320
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    • 2013
  • We studied the polarization state in an apertureless scanning near-field microscopy (a-SNOM) operating in reflection mode by using three-dimensional Finite-difference Time-domain (FDTD) method. As a result, the electric field around tip apex in the near-field region enhanced four times stronger than the incident light for ppolarization when the tip-sample separation was 10 nm. We find that the p- and s-polarization state is maintained for the scattered light when the probe is perpendicular to the sample. When the probe is not perpendicular to the sample, the polarization state of scattered light will rotate an angle that equals to the inclination angle of probe with p-polarization illumination. On the other hand, the polarization state will not rotate with s-polarization illumination.