Two-photon polymerization and crosslinking are commonly used methods for microfabrication of three-dimensional structures with applications spanning from photonic microdevices, drug delivery systems, to cellular scaffolds. However, the use of two-photon processes for precise, internal modification of biological tissues has not yet been reported. One of the major challenges has been a lack of appropriate tools to monitor and characterize crosslinked regions nondestructively.
Here, we demonstrate spatially selective two-photon collagen crosslinking (2P-CXL) in intact tissue for the first time. Using riboflavin photosensitizer and femtosecond laser irradiation, we crosslinked a small volume of tissue within animal corneas. Collagen fiber orientations and photobleaching were characterized by second harmonic generation and two-photon fluorescence imaging, respectively. Using confocal Brillouin microscopy, we measured local changes in longitudinal mechanical moduli and visualized the cross-linked pattern without perturbing surrounding non-irradiated regions. 2P-CXL-induced tissue stiffening was comparable to that achieved with conventional one-photon CXL. Our results demonstrate the ability to selectively stiffen biological tissue in situ at high spatial resolution, with broad implications in ophthalmology, laser surgery, and tissue engineering.
Conventional light-activated therapies, such as photodynamic therapy (PDT), photochemical tissue bonding (PTB), collagen crosslinking (CXL), low-level light therapy (LLLT), and antimicrobial therapy utilize external light sources and light propagation through free space, limiting treatment to accessible and superficial areas of the body. Recent progress has been made in developing biocompatible polymer waveguides to enhance light delivery to deep tissues. To further expand clinical utility, waveguides should be flexible and tough enough to enable use in anatomically difficult-to-reach regions, while having the requisite optical properties to achieve uniform and efficient illumination of the target area. Here, we present a new class of flexible polymer waveguides optimized for uniform light extraction into tissues. Our slab waveguides comprise two designs: first, a flexible polydimethylsiloxane (PDMS) based elastomer for CXL, and second, a tough polyacrylamide and alginate hydrogel for large-area phototherapies. Our waveguides are optically transparent in the visible wavelengths (400-750 nm) and a multimode fiber is used to couple light into the waveguide. We characterized the light propagation through the waveguides and light extraction into tissue, and validated our results with optical simulation. By changing the thickness and scattering properties, uniform light extraction through the length of the waveguide could be achieved. We demonstrate proof-of-concept scleral photo-crosslinking of an ex vivo porcine eyeball for prevention of myopia.
When waves travel through disordered media such as ground glass and skin tissues, they are scattered multiple times. Most of the incoming energy bounces back at the superficial layers and only a small fraction can penetrate deep inside. This has been a limiting factor for the working depth of various optical techniques. We present a systematic method to enhance wave penetration to the scattering media. Specifically, we measured the reflection matrix of a disordered medium with wide angular coverage for each orthogonal polarization states. From the reflection matrix, we identified reflection eigenchannels of the medium, and shaped the incident wave into the reflection eigenchannel with smallest eigenvalue, which we call anti-reflection mode. This makes reflectance reduced and wave penetration increased as a result of the energy conservation. We demonstrated transmission enhancement by more than a factor of 3 by the coupling of the incident waves to the anti-reflection modes. Based on the uneven distribution of eigenvalues of reflection eigenchannels, we further developed an iterative feedback control method for finding and coupling light to anti-reflection modes. Since this adaptive control method can keep up with sample perturbation, it promotes the applicability of exploiting reflection eigenchannels. Our approach of delivering light deep into the scattering media will contribute to enhancing the sensitivity of detecting objects hidden under scattering layers, which is universal problem ranging from geology to life science.
We implement differential interference contrast (DIC) microscopy using high-speed synthetic aperture imaging that expands the passband of coherent imaging by a factor of 2.2. For an aperture synthesized coherent image, we apply for the numerical post-processing and obtain a high-contrast DIC image for arbitrary shearing direction and bias retardation. In addition, we obtain images at different depths without a scanning objective lens by numerically propagating the acquired coherent images. Our method achieves high-resolution and high-contrast 3-D DIC imaging of live biological cells. The proposed method will be useful for monitoring 3-D dynamics of intracellular particles.
KEYWORDS: Charge-coupled devices, Molybdenum, Digital holography, Holography, Holograms, Microscopes, 3D image reconstruction, 3D metrology, Optical engineering, 3D image processing
Holography has a considerable advantage of retrieving three-dimensional information of an object from only one interference recording. However, twin images always appear in the reconstruction for the reason of symmetry. Especially, twin images significantly deteriorate the quality of the reconstructed information in on-axis configuration. A solution of the twin-image problem in a digital holographic microscope by using symmetry with quadrantal masks is suggested in this study. This method is effective to most of the measured area without any additional implements, and restrictions on sample or iterations, and is demonstrated by the simulation and experimental results. The ratio of the disturbed area by a twin-image to the total measured area is reduced to the value of 0.82% in a specific case.
We study the tolerance characteristic of SIAX and suggest a newly designed SIL-Axicon system for the better tolerances.
Methods for checking beam quality, optimization and remaining problems are suggested. SIL-Axicon system shows
more tolerances in the uniformity of beam incident angle. Bessel beam (BB) with SIL can be used for multi layer high
density data storage systems. We study the tolerance characteristic of SIAX and suggest a newly designed SIL-Axicon
system for the better tolerances.
Digital holography (DH) has a big advantage to retrieve the
three-dimensional (3D) information of the object from only
one interference recording. Especially, the digital holographic microscope (DHM) using a microscope objective (MO)
has been researched for 3D microscopy. The researches have progressed for compensation of aberrations and
improvement of the resolution in the optical system in recent years. Most of small aberrations caused by a MO are
compensated through various researches. However, the measured phase is distorted in the optical system, which has the
significant wavefront deformation in illuminating wave larger than number of wavelengths. In this paper, the relation
between illuminating wave and the reconstructed phase is studied based on the wave optics and the analysis is confirmed
by the simulations. The analysis of the wavefront compensation is applied to a super-resolution DHM in theory and the
technique for retrieving the distribution of the intensity and phase is demonstrated in simulation.
The digital holographic microscope (DHM) has emerged as a useful tool for verifying the three-dimensional structure of an object. A dual-type inline DHM that can be used with both transmission and reflection imaging in a single device is developed. The proper modes (between transmission and reflection imaging) can be easily changed according to the characteristics of the object in this system. The optimum condition for retrieving the correct phase information is illuminating a plane wave to an object. In contrast to the transmission imaging, it is difficult to illuminate an object using a plane wave without deformations in the reflection imaging. We developed an adequate relay lens module for illumination that can be adapted to any type of microscope objective without significant aberrations in the reflection imaging. The relationship between the illuminating condition and the measured phase based on the wave optics is analyzed. A specially designed and manufactured phase mask is observed in this system, and an alternative method for overcoming the limitation of phase unwrapping is introduced for the inspection of that object.
Holography has a considerable advantage to retrieve the three-dimensional (3D) information of an object from only one
interference recording. For several decades, the technology of digital holography (DH), which uses numerical
reconstruction as opposed to illuminating the reference beam to the hologram plate, has progressed with the assistance of
improvements in 2D array detectors and computers. In this paper, a dual-type inline digital hologram microscope (DHM)
system that can be used with both transmission imaging and reflection imaging in a single device is developed. The
proper method depending on the modes (transmission imaging or reflection imaging) can be changed easily in this
system according to the characteristics of the object. Illumination with a plane wave is the necessary condition for
retrieving the correct phase information. In the case of reflection imaging, unlike in transmission imaging, a special relay
lens in addition to the microscope objectives (MOs) is needed to meet the needs of this condition. However, the quality
of the 3D information can deteriorate significantly due to the overlapping twin image that is inherent in holography. This
study suggests an effective and convenient method for eliminating the twin image that is entangled in the reconstructed
information. The proposed method does not require extra components, numerical iterations, and restrictions on the
object.
According to the ITRS roadmap, DRAM half pitch (hp) will reach to 32 and 20 nm in 2012 and 2017
respectively. However, it is difficult to make sub-40 nm node by single exposure technology with currently
available 1.35 numerical aperture (NA) ArF immersion lithography. Although it is expected to enable 32 nm hp
with either double patterning technology or extreme ultra-violet lithography, there are many problems to be
solved with cost reduction. Thus, the study of high-index fluid immersion technology should be pursued
simultaneously. ArF water immersion systems with 1.35 NA have already introduced for 40 nm hp production.
ArF immersion lithography using high-index materials is being researched for the next generation lithography.
Currently, many studies are undergoing in order to increase NA with higher index fluid and lens in immersion
technology. The combination of LuAG (n=2.14) and third-generation fluid could be used to make 1.55 NA. This
combination with 0.25 k1, 32 nm hp can be obtained by single exposure technology. In order to check the
realization of this process and to check the possible process hurdles for this high NA single exposure technology,
32 nm hp with 1:1 line and space patterning is tried. Various illumination conditions are tried to make 1:1 32 nm
hp and the exposure and develop conditions are varied to check whether this single exposure can give
processible window. As a result, 32 nm hp can be obtained by single exposure technology with 1.55 NA.
Making a sub-100 nm contact hole pattern is one of the difficult issues in semiconductor process. Compared with
another fabrication process, resist reflow process is a good method to obtain very high resolution contact hole. However
it is not easy to predict the actual reflow result by simulation because very complex physics and/or chemistry are
involved in resist reflow process. We must know accurate physical and chemical constant values and many fabrication
variables for better prediction. We made resist reflow simulation tool to predict approximate resist reflow as functions of
pitch, temperature, time, array, and so on. We were able to see the simulated top view, side view and the changed hole
size. We used Navier-Stokes equation for resist reflow. We had varied the reflow time, temperature, surface tension, and
3-dimensional volume effect for old model. However the photoresist adhesion is another very important factor that was
not included in the old model. So the adhesion effect was added on Navier-Stokes equation and found that there was a
distinctive difference in reflowed resist profile and the contact hole width compared to the case of no adhesion effect.
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