Visualization Materials Using Silicon‐Based Optical Nanodisks (ViSiON) for Enhanced NIR Imaging in Ophthalmology

Abstract

ViSiON (visualization materials composed of silicon‐based optical nanodisks) is presented, which offers a unique optical combination of near‐infrared (NIR) optical properties and biodegradability. Initially, numerical simulations are conducted to calculate the total extinction and scattering effects of ViSiON by the diameter‐to‐thickness ratio, predicting precise control over its scattering properties in the NIR region. A top‐down patterning technique is employed to synthesize ViSiON with accurate diameter and thickness control. ViSiON with a 50 nm thickness exhibits scattering properties over 400 times higher than that of 30 nm, rendering it suitable as a contrast agent for optical coherence tomography (OCT), especially in ophthalmic applications. Furthermore, ViSiON possesses inherent biodegradability in media, with ≈95% degradation occurring after 48 h, and the degradation rate can be finely tuned based on the quantity of protein coating applied to the surface. Subsequently, the OCT imaging capability is validated even within vessels smaller than 300 µm, simulating retinal vasculature using a retinal phantom. Then, using an ex ovo chick embryo model, it is demonstrated that ViSiON enhances the strength of protein membranes by 6.17 times, thereby presenting the potential for ViSiON as an OCT imaging probe capable of diagnosing retinal diseases.

ViSiON, a novel visualization material composed of silicon‐based optical nanodisks, combines unique near‐infrared optical properties with biodegradability. Through numerical simulations and top‐down patterning, precise control over ViSiON's scattering properties is achieved, making it a promising contrast agent for optical coherence tomography, especially in ophthalmic applications.

1. Introduction

Optical coherence tomography (OCT) is a noninvasive and noncontact imaging technique that employs light wave reflection for high‐resolution cross‐sectional imaging of deep tissues.^{[ 1} , ^{2 ]} It is a rapidly advancing tool for observing tissue physiology and functionality, offering a means to enhance the visualization of pathological conditions. OCT has demonstrated its significance as a valuable bio‐imaging method in clinical applications including ophthalmology, cardiology, gastroenterology, and dermatology. However, its limitations include restricted contrast and limited penetration depth.^{[ 3} , ^{4 ]}

To improve imaging capability, several plasmonic nanomaterials, such as gold, iron oxide, silica, and polymer, have been proposed as contrast agents.^{[ 5} , ⁶ , ⁷ , ⁸ , ^{9 ]} But while nanotechnology can be utilized as a valuable tool for highly sensitive and accurate imaging, safety concerns should be addressed for successful clinical translations.^{[ 10 ]} To date, long‐term toxicity assessments of new materials have remained challenging and ambiguous, and further, predicting the side effects caused by nanomaterials remaining in the body after completing their role is also difficult.^{[ 11} , ^{12 ]} To overcome these issues, the development of nanomaterials that biodegrade after circulation in the body for a certain period is considered an ideal approach.^{[ 13} , ¹⁴ , ^{15 ]}

Among various nanomaterials, silicon‐based materials are generally recognized to have excellent biodegradability and unique intrinsic optical properties, and thus many studies have been conducted to apply them as contrast agents for OCT, as well as photoacoustic imaging, another powerful and noninvasive real‐time imaging tool.^{[ 16} , ¹⁷ , ^{18 ]} However, unlike other nanoparticles, controlling the size and shape of silicon‐based nanomaterials to optimize their performance as contrast agents has proven difficult.^{[ 19} , ²⁰ , ^{21 ]} It is therefore necessary to develop a synthetic strategy for the fabrication of biodegradable silicon nanomaterials with facile dimensional control within the near‐infrared (NIR) biological imaging window.

Here, we introduce ViSiON—visualization materials composed of silicon‐based optical nanodisks—with precisely tuned dimensions in the NIR region for examining blood vessels of the eye (Scheme  1 ). From numerical simulations, various candidate nanostructures were first designed and compared based on the fact that the total extinction and scattering cross‐sections can be controlled by changing the diameter/thickness ratio of the disks. We then synthesized intricate disk‐shaped nanoparticles with a specific diameter and thickness using a top‐down nanopatterning approach and compared them with the simulation results (Scheme 1a). The best candidates were evaluated in terms of their performance as contrast agents for OCT, which is commonly applied in ophthalmic examinations focusing on the NIR region.

Scheme 1.

a) Workflow for fabricating ViSiON through a top‐down nanopatterning technique incorporating simulations that analyze optical properties based on various nanodisk diameters and thicknesses. b) Optical coherence tomography (OCT) signal measurements inside a retina‐mimicking phantom utilizing ViSiON with scattering, as well as biodegradable properties.

Next, the biodegradability of ViSiON was confirmed in biological media along with the possibility of controlling the biodegradation kinetics by coating the disk surfaces with bovine serum albumin (BSA) (Scheme 1b). Two experiments were then conducted to test the applicability of ViSiON to the imaging of blood vessels and protein membranes. Application to a retina phantom model indicated that the high OCT signal intensity produced by ViSiON has potential for the imaging of blood vessels. Then, an in vivo experiment was performed with an ex ovo chick embryo model, which is useful for the study of ocular diseases and retinal development based on its similarity in anatomical structure to the human eye,^{[ 22} , ^{23 ]} demonstrating the capability of ViSiON to image protein membranes.

2. Results and Discussion

2.1. Simulated Optical Property Characterization According to ViSiON Diameter and Thickness

In recent years, high‐refractive‐index dielectric nanoparticles, such as silicon nanodisks, have attracted increasing attention due to the unique light–matter interaction associated with their electric and magnetic dipole or multi‐pole Mie resonances.^{[ 24} , ²⁵ , ^{26 ]} Through an alternative mechanism differing from that of metallic nanoparticles based on localized surface plasmon resonance, we expect that high‐refractive‐index silicon nanodisks can interact with incident light in the visible and NIR spectral ranges and thus ultimately be used as contrast agents for OCT. Accordingly, we designed ViSiON, short for visualization materials composed of silicon‐based optical nanodisks, for an OCT contrast agent.

As a first step to assess this potential, the extinction, scattering, and absorption cross sections of ViSiON with a nanodisk thickness of 30 nm and different diameters from 100 to 300 nm were numerically calculated (Figure  1a and Figure S1, Supporting Information). Figure 1a shows that the scattering cross‐sectional area in the visible and NIR regions increases as the diameter of ViSiON increases. In particular, the two peaks and two dips of the scattering spectra for 300 nm clearly show the characteristic features of high‐refractive‐index dielectric nanostructures.^{[ 24} , ²⁵ , ²⁶ , ²⁷ , ^{28 ]} The two peaks are known to arise from electric and magnetic dipole interactions, and the two dips are due to radiation‐less anapole states.^{[ 24} , ²⁷ , ^{28 ]} These two peaks and dips were redshifted with increasing ViSiON diameter and did not appear when the diameter‐to‐height ratio was low such as 100 nm. Both of these results are in good agreement with previous reports.^{[ 27} , ^{28 ]}

Figure 1.

Numerically simulated scattering spectra of ViSiON a) with a thickness of 30 nm and various diameters (100, 200, 250, and 300 nm), and b) with a diameter of 300 nm and various thicknesses (30, 40, and 50 nm). To simulate the use of ViSiON as contrast agents, a medium with a constant refractive index of 1.33 was assumed. c) Experimental absorbance spectrum of 300 nm ViSiON with thicknesses of 30, 40, and 50 nm. Inset: photograph of corresponding ViSiON solutions. d) Scanning electron microscope (SEM) image of 300 nm ViSiON with 50 nm thickness.

To investigate a possible route to further increase the ViSiON scattering efficiency at a wavelength of 850 nm, which is the center wavelength of the OCT light source used in this work, the scattering cross‐section of the 300 nm diameter ViSiON was calculated while varying the nanodisk thickness from 30 to 50 nm. The scattering cross sections of the ViSiON at a wavelength of 850 nm according to changes in thickness are summarized in Figure 1b. Compared to the nanodisks with 30 nm thickness and 100 nm diameter, ViSiON with 50 nm thickness and 300 nm diameter shows an almost 400 times larger scattering cross‐section, which is about 10 500 nm². This value is similar to the scattering cross‐section of a 200 nm diameter gold nanoparticle (about 12 000 nm²).^{[ 29 ]}

2.2. Experimental Preparation and Characterization of ViSiON

To synthesize the ViSiON as designed by numerical simulation, we applied nanoimprinting lithography and thermal evaporation, methods that can be found in previous reports (Figure S3, Supporting Information).^{[ 30 ]} Scanning electron microscope (SEM) images of synthesized ViSiON on Al (1% Si)‐coated Si wafers before release are shown in Figure S4a,b, Supporting Information. The pattern period and diameter were calculated as 600 and 300 nm, respectively, and the SEM image in Figure S4c, Supporting Information, indicates that the ViSiON had a high degree of uniformity.

We then synthesized 300 nm ViSiON with 30, 40, and 50 nm nanodisk thicknesses by controlling the amount of Si evaporation. The absorbance spectrum peaks shifted to the NIR region according to increasing thickness, a result that well matches the simulation data in Figure 1c. Accordingly, we confirmed through the simulations and experiments that ViSiON with 50 nm thickness shows optimal optical properties in terms of the highest absorption over the whole measured visible and NIR wavelength range.

The SEM images in Figure 1d and Figure S5, Supporting Information, were taken after drying aliquots of the respective ViSiON solutions. Even after detachment from the wafer, the particles retained their uniform morphology and exhibited precisely controlled thickness.

2.3. In Vitro OCT Imaging with Various ViSiON

We applied ViSiON as a contrast agent for OCT by taking advantage of their high scattering efficiency in the NIR region. In vitro OCT imaging was performed with an array of glass capillary tubes to examine the performance of ViSiON as an OCT contrast agent in the NIR window. A spectral‐domain OCT system with a center wavelength of 850 nm and bandwidth of 100 nm was used as the light source, and different ViSiON samples were diluted to half concentrations from 0 to 150 pm (Figure S6a, Supporting Information). Figure S6b, Supporting Information, shows the cross‐sectional OCT imaging results of the glass tubes filled with ViSiON at different concentrations and thicknesses. Under the 50 nm thickness condition, a higher OCT signal was generated compared to the other solutions at the same concentration. Here, the OCT intensity decreases toward the bottom of the glass tube, a phenomenon that occurs because the light is strongly scattered in the upper area and thus the light transmitted to the lower area decreases. The mean OCT signal intensities were measured with ImageJ, and the resulting graph shows that ViSiON with 50 nm thickness had the highest OCT signal at all concentrations (Figure S6c, Supporting Information). The OCT intensities of ViSiON with 50 nm thickness increased by 1.15 and 1.12 times compared to the 30 and 40 nm thickness groups, respectively, due to the shadow effect in the high concentration range (150–300 pm), whereas it increased by 1.57 and 1.31 times in the low concentration range (19–37 pm). Therefore, further experiments analyzing OCT characteristics were conducted using ViSiON with a uniform nanodisk thickness of 50 nm.

As shown in Figure  2 , the penetration depths of ViSiON were examined using a layer of PDMS mixture adjusted to 0.8 wt% TiO₂ particles corresponding to the effect of the retinal nerve fiber layer. For this measurement, PDMS layers with various thicknesses (0, 50, 100, 200, and 300 µm) were stacked on top of square glass capillary tubes with 100 µm walls (Figure 2a). The cross‐sectional OCT imaging results show much stronger OCT intensities in the absence of the PDMS layer, and also that the OCT signal of ViSiON is maintained at all layer thicknesses, demonstrating its penetration depth (Figure 2b).

Figure 2.

a) Diagram of the experimental design to determine the penetration depths of ViSiON. b) Cross‐sectional OCT images of ViSiON at various concentrations of 0, 19, 37, 75, 150, and 300 pm. The white box in the upper left marks the outline of the glass capillary tube, and the yellow box marks the inner surface of the glass capillary tube. c) Graph of the mean value of OCT intensity with different concentrations under 300 µm PDMS thickness. (***p < 0.001).

In Figure S7, Supporting Information, the mean OCT intensities of ViSiON are plotted for different PDMS layer thicknesses. Although the OCT signal was greatly reduced at the 300 µm depth, it was 2.23 times higher than the blank at 300 pm concentration and 1.14 times higher at 19 pm, indicating a significant signal (Figure 2c). The mean thickness of 10 intra‐retinal layers is known to be 280 µm, and moreover, the ganglion cell layer, inner plexiform layer, and inner nuclear layer, which are regions where diseases frequently occur, have been measured to be 25, 60, and 88 nm, respectively.^{[ 31 ]} Accordingly, the level of sensitivity ViSiON demonstrated here is assumed to be suitable for retinal disease imaging.

2.4. Biodegradability of Bovine Serum Albumin‐Coated ViSiON (cViSiON)

Silicon materials are particularly attractive as contrast agent nanomaterials for use in the body owing to their biocompatibility, non‐toxicity, and tunable hydrolytic degradation in biorelevant media (Figure  3a). Here, we explored whether the biodegradation kinetics can be controlled by coating the ViSiON surfaces with BSA (cViSiON), which is known for its ease of physisorption and stability within the short observation time required for imaging applications. First, to probe the biodegradability of bare (uncoated) ViSiON, Cy3 was adsorbed on the ViSiON surface, followed by incubation in distilled water (DW) and phosphate‐buffered saline (PBS) (Figure 3b). It was observed that the ViSiON surface was dispersed in PBS, which might accelerate the degradation rate of the silicon materials compared to DW, and fluorescence increased as the adsorbed Cy3 was released. When comparing the fluorescence signals at 1 h, it was observed that the fluorescence signal of ViSiON, solubilized in PBS, increased by 38.2 times compared to the condition where it was solubilized in DW. Moreover, the biodegradability of ViSiON can be controlled by coating its surface with proteins, thus allowing modulation of the degradation rate. We coated ViSiON with various concentrations of BSA (0, 0.3, 3, and 30 mg mL⁻¹) and observed changes in the peaks at 500 and 780 nm in the absorbance spectra (Figure 3c). In the case of the 0 mg mL⁻¹ BSA concentration, the absorbance intensity decreased to 5.99% at 500 nm and 4.72% at 780 nm over time. ViSiON without BSA coating reduced by over half in peak values at 500 and 780 nm, measuring 51.1% and 52.7%, respectively, at 1 h, indicating the degradation of more than half of the ViSiON. After 48 h, the peaks at 500 and 780 nm decreased to 5.9% and 4.7%, respectively, signifying the degradation of most of the ViSiON. Comparing the data at 48 h, the biodegradable protective effect at BSA concentrations of 0.3 and 3 mg mL⁻¹ was observed to be 37.6% and 49.6% at 500 nm and 38.6% and 51.9% at 780 nm, respectively. However, under conditions with 30 mg mL⁻¹ BSA, it was observed that the biodegradable effect at 1 h was nearly absent, with values of 107% at 500 nm and 105% at 780 nm. Furthermore, after 48 h, the peaks at 500 and 780 nm decreased to 92.8%, indicating minimal degradation. We also confirmed the biodegradability of ViSiON and the biodegradation inhibitory effect of BSA via OCT (Figure 3d). Results show that the lower the BSA concentration, the higher the ViSiON resolution, which means that the scattering ability of the particles is reduced. When a glass tube was used, the OCT intensity decreased to 75.7% when the ViSiON was synthesized with 30 mg mL⁻¹ of BSA, 40% when synthesized with 3 mg mL⁻¹ of BSA, and 16.9% when not coated with BSA. Moreover, OCT cross‐sectional images according to incubation time showed decomposition even in the case of incubation in PBS for 30 min in the absence of BSA (Figure 3e). Through these results, we demonstrated that the synthesized ViSiON is biodegradable and that the degradation rate can be controlled by surface modification.

Figure 3.

a) Schematic representation comparing the biodegradability of bovine serum albumin (BSA)‐coated ViSiON (cViSiON) and bare ViSiON. b) Schematic and fluorescence graph of Cy3 release from ViSiON in DW and PBS. c) Absorbance spectra reflecting the stability of ViSiON in PBS at BSA concentrations of 0, 0.3, 3, and 30 mg mL⁻¹. d) OCT images showing the stability of ViSiON in PBS at BSA concentrations of 0, 0.3, 3, and 30 mg mL⁻¹. e) Time‐dependent biodegradable OCT images of cViSiON and bare ViSiON.

2.5. In Vitro and In Vivo Applications for OCT Imaging of cViSiON

Next, we performed OCT imaging in a retinal phantom to confirm the potential of cViSiON as a contrast agent for in vivo imaging. The retinal phantom used in the experiment was fabricated with microfluidic channels mimicking retinal blood vessels, as shown in Figure  4a and Figure S8, Supporting Information. The fabricated retinal phantom was designed with a 300 µm thick 0.8 wt% PDMS–TiO₂ top layer. Figure 4b shows en face and cross‐sectional OCT imaging results of the phantom without (top) and with (bottom) cViSiON. The en face images show that the OCT intensity in the retinal phantom increased by 3.5 times in the presence of cViSiON compared to the control group. Figure S9, Supporting Information, shows the OCT signal differences after the injection of cViSiON at concentrations of 0, 50, 200, and 400 pm within a microfluidic channel mimicking retinal blood vessels. The images of the yz‐ and xz‐planes show a clear OCT intensity in the channels under the 300 µm PDMS layer.

Figure 4.

a) Design of the retinal phantom based on microfluidic channels mimicking retinal blood vessels. b) In vitro OCT imaging application of cViSiON. Cross‐sectional OCT images of the phantom without (top) and with (bottom) cViSiON are shown. The scale bars are 500 µm (middle) and 200 µm (right).

Finally, to demonstrate the potential utility of ViSiON in the field of retinal disease imaging, we conducted in vivo experiments using a chick embryo chorioallantoic membrane (CAM) model (Figure  5a). For this purpose, we utilized CAM that had matured for over 5 days, primarily because starting from the 4th day, undifferentiated microvessels became evident in all CAM blood vessels.^{[ 32 ]} Figure S10, Supporting Information, presents images of the setup for measuring OCT signals of eggs matured for 5 days with the eggshell removed. The OCT B‐scan images in Figure 5b are obtained after removing the eggshell and show cross‐sectional images both before (top) and after (bottom) the injection of cViSiON into the protein membrane, along with 3D reconstructions (right). ViSiON substantially enhanced the OCT signal intensity within the blood vessels by a factor of 6.17, resulting in clear imaging. The results from these in vivo imaging experiments confirm the potential applicability of ViSiON for tissue tomography.

Figure 5.

Ex ovo application of cViSiON for OCT imaging. a) Ex ovo model using a chick chorioallantoic membrane model similar to human retinal blood vessels. b) Cross‐sectional and projective views of OCT images of a 3‐day‐old developed chick embryo before and after injection of cViSiON. The labels I.M., B.V., C.E., and A.E. represent the inner membrane, blood vessels, chorionic epithelium, and allantoic epithelium, respectively. The scale bars are all 500 µm. The color bar for OCT is in logarithmic scale.

This study has two distinctive features. First, it employed top‐down patterning to control the thickness and diameter of silicon nanoparticles, thereby adjusting their scattering characteristics. Previous research on the scattering properties of silicon nanoparticles has focused on patterning silicon on substrates for applications in electronic components, solar cells, and optical devices. In contrast, the silicon nanodisks in our approach are released from the substrate for utilization as contrast agents in biological imaging, enabling the successful synthesis of highly uniform silicon nanoparticles.

Second, this study is positioned at the forefront in utilizing silicon nanoparticles as contrast agents for OCT. Recent studies have demonstrated active development in contrast agents utilizing the optical properties of silicon nanoparticles. Silicon nanoparticles have been employed as contrast agents for MRI,^{[ 33 ]} and technologies, such as fluorescence‐based imaging,^{[ 34 ]} have been developed using silicon nanoparticles with high fluorescence quantum yield. However, as of now, no published report has combined biodegradability with a fine‐tuning of the scattering properties of silicon nanoparticles for use as OCT contrast agents, making our paper distinctive in this aspect.

3. Conclusion

We presented visualization materials composed of silicon‐based optical nanodisks, or ViSiON, which provide unique optical properties in the NIR region and controllable biodegradation in biological media. Through numerical simulations, we first confirmed that the total extinction and scattering cross‐sections can be changed depending on the diameter/thickness ratio of the nanodisks. Based on the simulation results, ViSiON with an optimal diameter/thickness ratio was found to be suitable as a contrast agent for OCT based on their scattering properties in the NIR region. We demonstrated both biodegradability and the potential to control the biodegradation kinetics by coating the disk surfaces with BSA. Then the potential of ViSiON as an imaging probe was confirmed by performing blood vessel imaging through a retinal phantom that mimics retinal blood vessels and in vivo imaging through an ex ovo chick embryo model

4. Experimental Section

Materials

For the synthesis of silicon‐based nanodisks, the following materials, reagents, and equipment were used: 4‐in. aluminum (Al) sputtered Si wafers, a nanopillar‐patterned Si mold P600h‐p‐100d (Eulitha), Si pellets (99.99+%), PMGI‐SF3 solution (Kayaku Advanced Materials), mr‐I 8020R polymer (Micro Resist Technology), AZ 300 MIF developer (AZ Electronic Materials), Microdeposit Remover 1165 (Dow Electronic Materials), trisodium citrate dehydrate (99%, Sigma Aldrich), BSA (Sigma Aldrich), a spin coater (SPIN‐3000A, Midas), a nanoimprint lithography system (ANT‐4H, Exatech, Republic of Korea), and a UV spectrophotometer (UV‐2600, Shimadzu, Kyoto, Japan). For the fabrication of the retinal phantom, polydimethylsiloxane (PDMS, Sylgard 184 kit, Dow Corning, USA) and titanium dioxide (TiO2) powder (718 467, Sigma–Aldrich, USA) were used.

Synthesis of Visualization Materials Composed of Silicon‐Based Optical Nanodisks (ViSiON)

A 4‐in. Si wafer sputtered with Al of 100 nm thickness was prepared, and PMGI‐SF3 was spin‐coated on the wafer substrate at 3000 rpm for 30 min and then baked at 200 °C for 5 min for the sacrificial layer. After cooling down to room temperature, mr‐I 8020R was spin‐coated on the sacrificial layer and annealed for 1 min at 100 °C for lithography patterning. Subsequently, the nanopattern was thermally imprinted with a nanopillar‐patterned Si mold with a diameter of 300 nm for 3 min under a pressure of 30 bar at 170 °C. After nanoimprinting, reactive ion etching with O₂ plasma was conducted with 6 sccm of Ar gas and 10 sccm of O₂ gas at a working pressure of 70 mTorr and plasma RF power of 30 W, during which the mr‐I 8020R layer was etched. The PMGI‐SF3 layer was dissolved by wet etching using AZ 300 MIF developer for 5 s, exposing the Al layer surface on the Si wafer.

Silicon was deposited at thicknesses of 30, 40, and 50 nm using a thermal evaporator under vacuum conditions. After Si film deposition, the PMGI‐SF3 layer was removed in Microdeposit Remover 1165 solution under sonication for 20 min at 50 °C. The substrate with ViSiON was then soaked and sonicated in 1 wt% trisodium citrate solution for 20 min at 50 °C to remove the Al layer and release all ViSiON from the substrate. The ViSiON was washed with distilled water and concentrated by centrifugation at 13 000 rpm for 5 min. The morphology of ViSiON was observed via a field emission SEM (SU‐8000, Hitachi, Tokyo, Japan). The images were analyzed with ImageJ software to define the diameter and thickness of the disks. The optical properties, such as optical density and resonance wavelength, of the ViSiON solutions were measured with a UV spectrophotometer (UV‐2600, Shimadzu, Kyoto, Japan).

Optical Coherence Tomography Imaging

In the OCT imaging setup, a superluminescent diode (SLD) was utilized as the broadband light source. This particular SLD had a center wavelength of 849 nm (specifically, the BLM2‐D‐840‐B‐I‐10 model from Superlum, Ireland) and a −3 dB bandwidth spanning 100 nm. To distribute the light, it was equally divided into a reference arm and a sample arm through a 2 × 2 fiber optic coupler. Subsequently, the light emerging from the sample arm was directed onto the sample itself using a 2D galvanometer scanner and an objective lens. Finally, the reflected light from both the reference and sample arms was combined again and directed toward a linear wavenumber spectrometer.

Fabrication of the Retinal Phantom Model

A retinal phantom with a hexagonal pattern of four microfluidic channels that mimic blood vessels was fabricated. After making a mask with a hexagonal blood vessel design, a mold with a line width and height of 20 µm was made on a 4‐in. Si wafer through a lithography process. The phantom was completed by making transparent microfluidic channels using PDMS and the mold and then attaching a layer of 0.8% PDMS–TiO₂ mixture on the phantom. A 300 µm PDMS–TiO₂ layer thickness was obtained through spin coating. Fluorinated ethylene propylene (FEP) tubing was inserted into the inlet holes of the microchannels, and the fluid was injected with a syringe.

In Vivo Imaging Using the Ex Ovo Chick Embryo Model

For chick embryo imaging, fertilized chicken eggs cultivated at 37 °C and 45% humidity for 3 days were used. Only the upper portion of the eggshell was removed for OCT imaging, and the temperature was maintained at 37 °C during imaging using a hot plate. The OCT setup for the in vivo chick embryo imaging follows the setup mentioned above. The 3D volume was acquired with 512 A‐lines in a B‐scan, and the field of view was 4 mm × 4 mm with 3.2 µm axial resolution in air. The 3D images were reconstructed with commercial software (Amira 6, FEI, USA).

Conflict of Interest

The authors declare no conflict of interest.

Supporting information

Supporting Information

Ki J., Lee H., Lee T. G., Lee S.‐W., Wi J.‐S., Na H.‐K., Visualization Materials Using Silicon‐Based Optical Nanodisks (ViSiON) for Enhanced NIR Imaging in Ophthalmology. Adv. Healthcare Mater. 2024, 13, 2303713. 10.1002/adhm.202303713

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.