Search

Single-molecule localization microscopy (SMLM) surpasses the diffraction limit by randomly switching fluorophores between fluorescent and dark states, precisely pinpointing the resulted isolated emission patterns, thereby reconstructing the super-resolution images based on the accumulated locations of thousands to millions of single molecules. This technique achieves a ten-fold improvement in resolution, unveiling the intricate details of molecular activities and structures in cells and tissues. Multicolor SMLM extends this capability by imaging distinct protein species labeled with various fluorescent probes, providing insights into structural intricacies and spatial relationships among different targets. This review explores recent advancements in multicolor SMLM, evaluates the strengths and limitations of each variant, and discusses the future prospects. [ABSTRACT FROM AUTHOR]

This article presents a numerical analysis of a very thin concentric octagonal elliptical ring resonator (CORR) with a cylindrical rod optical nano-metamaterial absorber (ONMMA) that is designed to absorb solar energy in the infrared, visible, and ultraviolet ranges. The ONMMA is made of gold and nickel resonators on a quartz substrate with a gold plate ground material. The absorption rate of the ONMMA is over 90% from 593.84–829.28 THz, with an average absorption rate of 93.891%. This means that the ONMMA can absorb more than 80% of the energy in the infrared, visible, and a small portion of the ultraviolet range. It also has an absorption rate of over 75% from 226.64–853.04 THz, with an average absorption rate of 88.945%. This covers the entire visible spectrum, with a small portion in the infrared and ultraviolet regions. The design has an operational bandwidth of 200–920 THz, with an average absorption rate of 86.424%. The ONMMA achieves a maximum absorption rate of 99.866% at 705.44 THz. The design of the octagonal ONMMA ensures that it is not affected by the polarization of the incident light and can absorb energy even at oblique angles up to 60 degrees. The study also analyzed the effects of different materials and geometrical parameters on the absorption characteristics, as well as the distribution of surface current, magnetic field, electric field, and power loss. The proposed ONMMA has promising applications in infrared detection, solar cells, and multicolor imaging applications. [ABSTRACT FROM AUTHOR]

Abstract Single-molecule localization microscopy (SMLM) surpasses the diffraction limit by randomly switching fluorophores between fluorescent and dark states, precisely pinpointing the resulted isolated emission patterns, thereby reconstructing the super-resolution images based on the accumulated locations of thousands to millions of single molecules. This technique achieves a ten-fold improvement in resolution, unveiling the intricate details of molecular activities and structures in cells and tissues. Multicolor SMLM extends this capability by imaging distinct protein species labeled with various fluorescent probes, providing insights into structural intricacies and spatial relationships among different targets. This review explores recent advancements in multicolor SMLM, evaluates the strengths and limitations of each variant, and discusses the future prospects.

Purpose: To characterize features of central retinal artery occlusion (CRAO) using multicolor (MC) imaging and to assess the differences in CRAO grading between color fundus photography (CFP) and MC image qualitatively and quantitatively. Methods: We conducted a prospective, cross-sectional study in the Department of Ophthalmology of Renmin Hospital of Wuhan University. In total, 86 acute CRAO patients were included. Spectral-domain optical coherence tomography (SD-OCT), CFP, and MC examinations were taken at baseline. Based on the findings of these three examinations, CRAO was divided into three grades (incomplete, subtotal, and total). Based on OCT grading criteria, we qualitatively compared the ability of grading CRAO by CFP and MC. CRAO patient's visual acuity (VA) was obtained from the initial visit. The retinal thickness was measured by SD-OCT. Superficial capillary plexus (SCP) and deep capillary plexus (DCP) were obtained from optical coherence tomography angiography (OCTA) examinations. Quantitative data were compared across the three acute CRAO subgroups and against three examination findings. Results: MC image had significantly higher power of acute CRAO detection than CFP (P = 0.03). In the same group of CRAO patients, there was no significant difference in VA when comparing OCT with the MC grading system or with the CFP grading system (all P > 0.05). Significant differences in VA were found between the three CRAO subgroups only under MC grading (P = 0.016). In incomplete CRAO patients, significant differences were found in central fovea thickness (CFT) when comparing OCT with the CFP grading system (P = 0.019). In the same group of CRAO patients, there was no significant difference in retinal thickness when comparing OCT with the MC grading system (All P > 0.05). Significance differences in CFT (P < 0.001), innermost retinal layer (IMRL; P < 0.01), middle retinal layer (MRL; P < 0.001), and outer retinal layer (ORL; P = 0.021) were found between the three CRAO subgroups by MC grading. Vessel density of SCP showed a statistically increased as the severity of three CRAO subgroups (P = 0.03), whereas DCP did not have significant differences (P = 0.745). Comparisons were made between the OCT grading method and the MC and CFP grading methods; there is no significant difference in vessel density of SCP and DCP (All P > 0.05). Conclusion: The images obtained by MC are superior to those obtained by CFP in CRAO grading, retinal thickness, and vessel density measurement. MC imaging may be more capable of CRAO grading than OCT. We recommend MC imaging to determine CRAO severity to guide disease treatment and predict visual prognosis. [ABSTRACT FROM AUTHOR]

Fluorescence imaging is an indispensable method for studying biological processes non-invasively in cells and transparent organisms. Extension into the shortwave infrared (SWIR, 1000-2000 nm) region of the electromagnetic spectrum has allowed for imaging in mammals with unprecedented depth and resolution for optical imaging. In this review, we summarize recent advances in imaging technologies, dye scaffold modifications, and incorporation of these dyes into probes for SWIR imaging in mice. Finally, we offer an outlook on the future of SWIR detection in the field of chemical biology.

Carbon dots (CDs) have recently garnered considerable attention as remarkable fluorescent nanomaterials. However, the production of thermally stable CDs still remains a significant challenge as it requires a stable structure resistant to thermal oxidation and effective control of aggregation sites. Herein, we have developed a strategy for producing fluorescent nano-CDs with a stable structure and high thermal resistance properties using citrazinic acid and urea via solid-state carbonization. These nano-CDs exhibit excellent performance at high temperatures ranging from 10 to 90 °C. The core structure of nano-CDs is functionalized with various CO/CN nitrogen and oxygen groups, which enhance the structural integrity of the fluorescent nano-CDs and protect them against thermal oxidation and the aggregation effect. The thermal stability of the nano-CDs was evaluated using state-of-the-art techniques such as photoluminescent spectroscopy, UV–vis absorption spectroscopy, and decay lifetime measurements. Besides thermal resistance, nano-CDs exhibit green light emission with a maximum excitation of 400 nm and a peak quantum yield of 7.8%. These low-toxicity nano-CDs were successfully incubated with T-ca cancer cells, enabling stable multicolor imaging even at elevated temperatures (25–45 °C). [ABSTRACT FROM AUTHOR]

Adapting MRI to the field of molecular imaging is a challenge that continues to attract considerable attention. The detection of a given biomolecular analyte requires the availability of a molecular probe that is able to respond with a sufficient signal gap in order to overcome the low sensitivity of MRI. To this end, 19F MRI has yielded promising results due to its negligible endogenous background signals. Several small molecular probes have been reported; however, the low sensitivity of 19F MRI render the detection of biomarkers in vivo difficult, in view of their sub‐nanomolar concentration on site. Researchers have thus directed their attention toward larger chemical objects containing large numbers of 19F nuclei as smart platforms that allow a stimuli‐induced response. This article discusses the recent advances and future directions in the design of nano‐probes that respond to chemical or physical stimuli recognized as biomarkers for life and disease. [ABSTRACT FROM AUTHOR]

Bandgap engineering plays an important role in the regulation of the photophysical properties of semiconductor materials. Developing the facile and powerful strategy to achieve bandgap tunability is highly desired. Herein, the bandgap tunability of scandium microspheres prepared by the hydrothermal treatment of Sc3+ ions/l‐glutamine (Sc3+/Gln) complexes is achieved. The continuously decreased bandgap is directly attributed to the increase of carbon element and the decrease of scandium element in scandium microspheres controlled by the Gln ligand, which causes the red‐shifted room temperature phosphorescence from Sc/Gln‐0.15 (3.17 eV, green) to Sc/Gln‐0.25 (2.53 eV, yellow). Meanwhile, scandium microspheres have an excitation‐dependent emission ranged from green to red color, revealing their multi‐color imaging capability. Further increase of Gln ligand can further reduce the bandgap to 1.62 eV (Sc/Gln‐0.3), realizing the transition from photoluminescence to photothermal properties of scandium microspheres. Because of their superior optical properties, scandium microspheres can be well used for advanced anti‐counterfeiting and multicolor imaging. This new finding not only offers a deeper insight into the photophysical structure of rare earth semiconductor materials, but also provides a scalable method for the preparation of scandium microspheres with the desired bandgaps. [ABSTRACT FROM AUTHOR]

PurposeTo compare retinal pathology visualization in multispectral scanning laser ophthalmoscope imaging between the Spectralis and Optos devices.MethodsThis retrospective cross-sectional study included 42 eyes from 30 patients with age-related macular degeneration (19 eyes), diabetic retinopathy (10 eyes), and epiretinal membrane (13 eyes). All patients underwent retinal imaging with a color fundus camera (broad-spectrum white light), the Spectralis HRA-2 system (3-color monochromatic lasers), and the Optos P200 system (2-color monochromatic lasers). The Optos image was cropped to a similar size as the Spectralis image. Seven masked graders marked retinal pathologies in each image within a 5 × 5 grid that included the macula.ResultsThe average area with detected retinal pathology in all eyes was larger in the Spectralis images compared with Optos images (32.4% larger, P < 0.0001), mainly because of better visualization of epiretinal membrane and retinal hemorrhage. The average detection rate of age-related macular degeneration and diabetic retinopathy pathologies was similar across the three modalities, whereas epiretinal membrane detection rate was significantly higher in the Spectralis images.ConclusionSpectralis tricolor multispectral scanning laser ophthalmoscope imaging had higher rate of pathology detection primarily because of better epiretinal membrane and retinal hemorrhage visualization compared with Optos bicolor multispectral scanning laser ophthalmoscope imaging.

Astrocytic hamartoma is a benign glial tumor. It may be associated with tuberous sclerosis and can also be found incidentally on retinal examination as an isolated presentation. Here, we describe multimodal imaging characteristics of astrocytic hamartoma in a patient with retinitis pigmentosa. Spectral domain optical coherence tomography of both eyes showed moth-eaten optically empty spaces and hyperreflective dots along with foveal thinning. Multicolor image highlighted mulberry appearance of the lesion with green shift signifying elevated lesion. In infrared reflectance, lesion was hyporeflective with its margins well delineated. Green reflectance and blue reflectance highlighted calcification as multiple hyperreflective dots. Autofluorescence showed typical hyperautofluorescence.

Recent progress in ultrafast spectroscopy and semiconductor technology is enabling unique applications in screening, detection, and diagnostics in the Terahertz (T-ray) regime. The promise of efficaciously operation in this spectral region is tempered by the lack of devices that can spectrally analyze samples at sufficient temporal and spatial resolution. Real-time, multispectral T-ray (Mul-T) imaging is reported by designing and demonstrating hyperspectral metamaterial focal plane array (MM-FPA) interfaces allowing multiband (and individually tunable) responses without compromising on the pixel size. These MM-FPAs are fully compatible with existing microfabrication technologies and have low noise when operating in the ambient environment. When tested with a set of frequency switchable quantum cascade lasers (QCLs) for multicolor illumination, both MM-FPAs and QCLs can be tuned to operate at multiple discrete THz frequencies to match analyte "fingerprints." Versatile imaging capabilities are presented, including unambiguous identification of concealed substances with intrinsic and/or human-engineered THz characteristics as well as effective diagnosis of cancerous tissues without notable spectral signatures in the THz range, underscoring the utility of applying multispectral approaches in this compelling wavelength range for sensing/identification and medical imaging.

PurposeOur purpose was to compare the characteristics of the retinal and choroidal lesions including choroidal nevus, choroidal melanoma and congenital hypertrophy of the retina pigment epithelium using conventional color fundus photography (CFP) and multicolor imaging (MCI).MethodsThe paired images of patients with retinal or choroidal lesions were assessed for the visibility of lesion's border, halo and drusen using a grading scale (0-2). The area of the lesion was measured on both imaging modalities. The same grading was also done on the individual color channels of MCI for a further evaluation.ResultsThirty-three eyes of 33 patients were included. There were no significant differences in the mean border, drusen and halo visibility scores between the two imaging modalities (p = 0.12, p = 0.70, p = 0.35). However, the mean area of the lesion was significantly smaller on MCI than that on CFP (14.9±3.3 versus 18.7±3.4 mm2, p = 0.01).ConclusionThe appearance of choroidal and/ or retinal lesions on MCI may be different than that on CFP. Though MCI can provide similar information with CFP for the features of retinal and/ or choroidal lesions including border, halo and drusen; the infrared light reflection on MCI underestimates the extent of the choroidal lesion by 33%.

Rapid multicolor three-dimensional (3D) imaging for centimeter-scale specimens with subcellular resolution remains a challenging but captivating scientific pursuit. Here, we present a fast, cost-effective, and robust multicolor whole-organ 3D imaging method assisted with ultraviolet (UV) surface excitation and vibratomy-assisted sectioning, termed translational rapid ultraviolet-excited sectioning tomography (TRUST). With an inexpensive UV light-emitting diode (UV-LED) and a color camera, TRUST achieves widefield exogenous molecular-specific fluorescence and endogenous content-rich autofluorescence imaging simultaneously while preserving low system complexity and system cost. Formalin-fixed specimens are stained layer by layer along with serial mechanical sectioning to achieve automated 3D imaging with high staining uniformity and time efficiency. 3D models of all vital organs in wild-type C57BL/6 mice with the 3D structure of their internal components (e.g., vessel network, glomeruli, and nerve tracts) can be reconstructed after imaging with TRUST to demonstrate its fast, robust, and high-content multicolor 3D imaging capability. Moreover, its potential for developmental biology has also been validated by imaging entire mouse embryos (~2 days for the embryo at the embryonic day of 15). TRUST offers a fast and cost-effective approach for high-resolution whole-organ multicolor 3D imaging while relieving researchers from the heavy sample preparation workload.

Metallic nanoparticles, especially gold nanoparticles (AuNPs), have been widely used as bright optical probes for the observation and analysis of biomolecules. By continuously acquiring optical images of AuNPs bound to target molecules and analyzing their central coordinates, the behavior of a single biological molecule can be captured with high localization precision and high temporal resolution. This technique has been applied to a variety of biological studies, such as elucidating the operation mechanism of motor proteins that move forward at intervals as small as several nm, and the dynamics of lipid molecules that diffuse rapidly across biological membranes. In this review, I will focus on multicolor, high-speed, and high-precision optical imaging methods using metallic nanoparticles. These developments pave the way for a detailed understanding of the mechanisms of operation of tiny and complex biomolecules. [ABSTRACT FROM AUTHOR]

Multicolor fluorescence microscopy is an essential tool to visualize structures and dynamics in the life and materials sciences. However, the near-simultaneous acquisition of labels differing in excitation spectrum is difficult and renders such measurements prone to artifacts. We present a simple strategy to provide quasi-simultaneous fluorescence imaging with multiple excitation wavelengths by using an optical element to displace the sample image on the sensor at a rate that is much faster than the image acquisition rate and synchronizing this with the illumination. The emission elicited by the different wavelengths can then be encoded into the point-spread function of the imaging or visualized as multiple distinct images. In doing so, our approach can eliminate or mitigate artifacts caused by temporal aliasing in conventional sequential imaging. We demonstrate the use of our system to uncover hidden emissive states in single quantum dots and for the imaging of Ca 2+ signaling in neurons.

PurposeTo characterize features of retinal never fiber in Leber Hereditary Optic Neuropathy (LHON) using multicolor (MC) imaging and color fundus photography (CFP).MethodsNinety-two eyes of patients with LHON underwent MC imaging, optic disc spectral domain optical coherence tomography (SD-OCT), and CFP. Two independent observers graded RNFL visibility scores and two other experts determined never fiber bundle defects from four-quadrant readings. CFP, standard MC, infrared reflectance (IR), green reflectance (GR), blue reflectance (BR), and green-blue-enhanced (BG) imaging were compared.ResultsAgreement on never fiber bundle defects was substantial for CFP, standard MC, GR, BR, and BG images relative to IR. It was shown that BR (2.71 ± 0.55) had the best mean RNFL visibility score, BG (2.69 ± 0.52), GR (2.69 ± 0.56), standard MC (2.04 ± 0.79), CFP (1.80 ± 0.82), and IR (0.45 ± 0.59) followed. Agreement on temporal area defects was relatively improved. Youden's indices for CFP (78.21%), standard MC (84.48%), GR (90.92%), BR (89.64%), and BG (90.99%) indicated good detection of defects in the papillomacular bundle (PMB)/ high suspicion of patients with LHON, particularly for BG and GR. According to the proportion of never fiber bundle defects, standard MC, GR, BR, and BG can roughly determine the LHON clinical stage, especially in subacute and chronic stages, and standard MC is superior for patients with LHON of all stages. The stage judged by MC was consistent with the course inferred by pRNFL thickness.ConclusionAs an adjunct to SD-OCT, the MC image, particularly the GR and BG can delineate RNFL more effectively than CFP. The MC image may be a useful adjunct to OCT for detecting or monitoring never fiber bundle defects, providing inexpensive and rapid methods that can quickly identify patients with high suspicion of LHON.

Choroidal hamartoma, color fundus photography, multicolor imaging, retinal hamartoma, tuberous sclerosis complex Keywords: Choroidal hamartoma; color fundus photography; multicolor imaging; retinal hamartoma; tuberous sclerosis complex EN Choroidal hamartoma color fundus photography multicolor imaging retinal hamartoma tuberous sclerosis complex 2720 2724 5 07/15/22 20220701 NES 220701 Tuberous sclerosis complex (TSC) or Bourneville disease is a neurocutaneous (phacomatosis) disorder characterized by the development of multiple benign tumors of the embryonic ectoderm involving the skin, eyes, and nervous system. The choroidal hamartoma was barely visible on clinical examination or documented on color fundus photographs, and the multicolor image made the lesion more obviously visible. [Extracted from the article]

Introduction: We report a male who presented with acute visual defect and was diagnosed with urticarial vasculitis with recurrent branch retinal artery occlusion (BRAO) after systemic disease survey, fluorescein angiography (FA), and MultiColor imaging (MCI). Case report: A 47-year-old male with a history of urticarial vasculitis presented with visual defect OD. Fundus examination showed two foci of ischemic retinal whitening beneath the inferior arcade and above the superior arcade. MCI demonstrated a greenish tinge in the corresponding area. FA revealed segmental arteriolar staining and arterial occlusive changes. BRAO with retinal arteritis was diagnosed. Toxoplasma IgG was positive. Sulfamethoxazole 400mg plus trimethoprim 80mg was given. His vision worsened after 1-week of treatment. The established lesions improved, but new lesions occurred. Interferon-gamma release assay was positive but tuberculosis DNA qualitative amplification test of sputum was negative. Sputum acid-fast stain was positive and culture revealed nontuberculous mycobacteria. Left facial itching and reactive lymphadenopathy developed. Prednisolone and cyclophosphamide were started. The initial retinal artery lesions regained perfusion. Conclusions: Urticarial vasculitis with recurrent BRAO is an immune complex-mediated disease. Greenish-tinged occlusive lesions were noted from MCI with high resolution and contrast. MCI could be a valuable method for retinal vessel occlusive disease detection before FA and follow up.

AIM: To investigate the application of rapid diagnosis and classification of retinal artery occlusion(RAO)by multicolor imaging(MCI)with optical coherence tomography(OCT).METHODS: Totally 19 patients(19 eyes)who were diagnosed with RAO in our hospital were retrospectively analyzed. All of the patients were diagnosed of RAO after ophthalmologic examination, including 13 eyes of central retinal artery occlusion(CRAO), 5 eyes of branch retinal artery occlusion(BRVO)and 1 eye of RAO with sparing of cilioretinal artery. The best corrected visual acuity, slit lamp, OCT, MCI, FFA and visual field were performed on the patients, and the examination results and image data were analyzed.RESULTS: Totally 12(63%)eyes showed dullness or disappearance of pupil response to light, 16(84%)eyes showed total or partial paleness and edema in the posterior pole of retina, and 10(53%)eyes showed narrowing of retinal artery by slit-lamp examination. FFA examination was performed in 10 cases(53%). However 9 cases were not examined, including 6 cases with serious systemic diseases, 2 cases rejecting to be checked and 1 case with drug allergy. All the patients showed specific manifestations in both MCI and OCT examination, and the typical highly reflectivity of inner retinal layers in OCT formed tight one-to-one relationship with the green ischemic area in MCI.CONCLUSION: RAO can be diagnosed quickly by MCI combined with OCT, and their image characteristics can accurately identify the retinal ischemic areas, which can help for disease classification and prognosis.

Diagnostic investigation on pachychoroid spectrum disease (PSD) has been growing along with the rapid advancement of imaging technology. In optical coherence tomography (OCT)-based studies, choroidal thickness profile, luminal and stromal choroidal ratio, and abnormalities in the neurosensory retina have demonstrated various patterns in different clinical entities related to PSD. The emerging role of OCT angiography ()CTA) has been expanded to involve the quantitative analysis of the OCTA parameters in different clinical entities of PSD and to evaluate the choriocapillaris signal void and vessel density as indicators of choriocapillaris ischemia. OCTA has broadened our knowledge in characterization and assessment of both active and quiescent choroidal neovascularization and its association with treatment response. Recent studies using indocyanine green angiography have focused on the evaluation of choroidal vascular hyperpermeability and its relationship with other pachychoroid related features. Ultra-widefield indocyanine green angiography enables observation and characterization of peripheral choroidal pathologies and their associations with macular abnormalities. Multicolor imaging is an emerging modality with the capability to demonstrate early abnormalities in PSD. We summarize all investigations reflecting the new insights into the application of multimodal imaging for PSD and focus on novel findings observed in different clinical entities with each imaging modality. [ABSTRACT FROM AUTHOR]

A simple and convenient preparation of multicolor carbon dot (CD) solutions was proposed using neutral red as the carbon and nitrogen sources by a layering and extraction procedure. After poly-(vinylpyrrolidone) (PVP) as the cross-linking agent was doped into as-prepared solutions, red-, orange-, and yellow-emitting CD powders were finally fabricated with solid-state fluorescence quantum (QY) yields of 6.33, 16.5, and 43.6%, respectively. Through studies of surface functional groups and photoluminescence (PL) property compared with solution and powder states of CDs, the cross-linking effect emission (CEE) brought by PVP was confirmed in charge of the high solid-state QY of CD powders. In addition, the conjugate structure of neutral red and increasing hydrophobicity are believed to be responsible for the long-wavelength emission of CD powders. Notably, multicolor imaging of latent fingerprints (LFPs) was successfully achieved on complex background surfaces under 365 nm UV illumination. As a result, these carbon-based phosphors are promising in forensic investigation, bioimaging, and sensing fields. [ABSTRACT FROM AUTHOR]

Purpose: To compare multicolor imaging (MCI) with Optos color fundus photography (OCFP) for the evaluation of morphology and extent of preretinal membranes in diabetic tractional retinal detachments (TRD).Methods: In this retrospective study, 30 eyes with diabetic TRDs were imaged using the MCI feature of the Heidelberg Spectralis Spectral-domain optical coherence tomography (SD-OCT) and color photo using the Optos Daytona ultra-widefield fundus camera. Two investigators independently graded and determined the agreeability between the two modalities with respect to the extent of the TRD and preretinal membranes on the SD-OCT B-scan images.Results: The MCI provided better visualization of the attachments and traction points of the posterior hyaloid face and preretinal membranes and is comparable to the SD-OCT B-scan images. The inter-rater agreeability rates for OCFP had a Kappa (κ) value of 0.37, while the MCI had a κ value of 0.46. When comparing between images of different wavelengths, grading using infrared reflectance (IR) had a poor agreement (-0.04 ± 0.04) while green reflectance (GR) (0.46 ± 0.32) and blue reflectance (BR) (0.53 ± 0.19) had a moderate agreement. The composite MCI and GR images also had comparatively higher intraclass coefficient when compared to the OCFP (0.25 [-0.09-0.55]) and IR (-0.03 [-0.39-0.34]) images.Conclusion: MCI is more sensitive for determining the extent of TRDs and for the detection of secondary membranes when compared to OCFP, thus, aiding in better surgical planning. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the accuracy of MultiColor imaging (MC) compared to fluorescein angiography (FA) in detecting proliferative diabetic retinopathy (PDR) and associated diabetic retinopathy features. Methods: Fifty-nine eyes from 38 PDR patients were included. MC images were reviewed by 2 independent masked graders. A qualitative analysis based on the following features was performed: neovascular complexes (NVC), disc neovascularization (NVD), neovascularization elsewhere (NVE), microaneurysm (MA), intraretinal hemorrhage (IRH), vitreous hemorrhage (VH), preretinal hemorrhage (PRH), fibrosis, hard exudates (HE), epiretinal membrane (ERM), diabetic macular edema (DME), ischemia and laser spots (LS). Measures of diagnostic accuracy compared to FA were determined. Results: The sensitivity for the detection of NVC using MC was 95.1%, with a specificity of 40.0%, positive predictive value (PPV) of 92.9% and negative predictive value (NPV) of 50.0%. Sensitivity and specificity were higher in detecting NVD (88.9% and 76.9%) while NVE registered higher PPV (88.9%). MC was highly sensitive in detecting IRH, HE, ERM and LS (100%), MA (98.0%) and fibrosis (95.5%). Highest specificity was found for VH (100.0%), DME (100.0%), PRH (98.1%) and LS (89.5%). The area under the receiver-operating characteristic analysis of MC was excellent in NVD (0.83, 95% confidence interval (CI), 0.71–0.95, p < 0.001), IRH (0.89, 95% CI 0.74–1.00, p < 0.001), VH (0.81, 95% CI 0.60–1.00, p = 0.005) and PRH (0.89, 95% CI 0.68–1.00, p = 0.004) and outstanding in LS detection (0.95, 95% CI 0.87–1.00, p < 0.001). These results are likely due to the contrast and quality of the MC since better discrimination is enabled by the green wavelength. Conclusion: MC is useful in evaluation of PDR patients and can complement noninvasive imaging. MC detected some PDR features more accurately than FA such as NVD, IRH, VH, PRH, and LS. [ABSTRACT FROM AUTHOR]

Choroidal osteoma is a rare benign tumor which is found in the posterior pole of the eye. We herein describe multimodal imaging in two cases of choroidal osteoma. Fundus of our first case showed a yellowish-orange colored subretinal lesion at the posterior pole. Multicolor imaging highlighted the lesion with greenish hue. Infrared reflectance showed hyporeflectance. A dense echogenic plaque persisting in lower gain was noted on B-scan. FFA showed hyperfluorescence with corresponding hypocyanescence on ICG. EDI OCT showed an increase in choroidal thickness with elevated retinal pigment epithelium. The second case showed choroidal osteoma with active choroidal neovascular membrane that responded to intravitreal injection of Ranibizumab.

Over the last decade, tissue-clearing techniques have expanded the scale of volumetric fluorescence imaging of the brain, allowing for the comprehensive analysis of neuronal circuits at a millimeter scale. Multicolor imaging is particularly powerful for circuit tracing with fluorescence microscopy. However, multicolor imaging of large samples often suffers from chromatic aberration, where different excitation wavelengths of light have different focal points. In this study, we evaluated chromatic aberrations for representative objective lenses and a clearing agent with confocal microscopy and found that axial aberration is particularly problematic. Moreover, the axial chromatic aberrations were often depth-dependent. Therefore, we developed a program that is able to align depths for different fluorescence channels based on reference samples with fluorescent beads or data from guide stars within biological samples. We showed that this correction program can successfully correct chromatic aberrations found in confocal images of multicolor-labeled brain tissues. Our simple post hoc correction strategy is useful to obtain large-scale multicolor images of cleared tissues with minimal chromatic aberrations. [ABSTRACT FROM AUTHOR]

Purpose: The current study compares the ability of multicolor imaging (MCI) to detect the lesions of central serous chorioretinopathy against conventional color fundus photographs (CFP). Methods: It was a retrospective, observational case series of 93 eyes of 58 patients of central serous chorioretinopathy who underwent MCI and CFP. MCI and spectral-domain optical coherence tomography (SD-OCT) were performed using Spectralis SD-OCT system (HRA + OCT). CFP was obtained using FF 450 Plus fundus camera (Carl Zeiss Meditec, Jena, Germany). SD-OCT was considered gold standard for subretinal fluid (SRF) and retinal pigment epithelium detachment (PED). CFP was considered confirmatory investigation for fibrin and blue autofluorescence image (BAF) was considered gold standard to detect retinal pigment epithelium (RPE) atrophy. Results: CFP could detect SRF in 41 (44.1%) eyes. MCI detected SRF in 43 (46.2%) eyes. The sensitivity and specificity of MCI to detect SRF were 70.7% and 94.3%, respectively. PED was detected by CFP in 21 (22.6%) eyes and MCI in 27 (29%) eyes. The sensitivity and specificity of MCI to detect PED were 70% and 97.7% respectively. CFP could pick RPE atrophy in 52 (55.9%) eyes whereas MCI was picked it in 78 (83.9%) of eyes. Conclusion: Both MCI and CFP were inferior to a gold standard in identifying the SRF, PED, and RPE atrophy. However, MCI was better than CFP in comparison with gold standard for these clinical findings in CSC. Thus, MCI seems to be a more valuable imaging tool compared to CFP.

Over the last decade, tissue-clearing techniques have expanded the scale of volumetric fluorescence imaging of the brain, allowing for the comprehensive analysis of neuronal circuits at a millimeter scale. Multicolor imaging is particularly powerful for circuit tracing with fluorescence microscopy. However, multicolor imaging of large samples often suffers from chromatic aberration, where different excitation wavelengths of light have different focal points. In this study, we evaluated chromatic aberrations for representative objective lenses and a clearing agent with confocal microscopy and found that axial aberration is particularly problematic. Moreover, the axial chromatic aberrations were often depth-dependent. Therefore, we developed a program that is able to align depths for different fluorescence channels based on reference samples with fluorescent beads or data from guide stars within biological samples. We showed that this correction program can successfully correct chromatic aberrations found in confocal images of multicolor-labeled brain tissues. Our simple post hoc correction strategy is useful to obtain large-scale multicolor images of cleared tissues with minimal chromatic aberrations.

Aims: To quantify hard exudates (HEs) by multicolor imaging (MCI) and traditional color fundus photography (CFP) in diabetic macular edema (DME), and study their associations with serum lipid levels. Methods: Observational study. DME patients with HEs were recruited. The HE area and location both by MCI and CFP were measured by ImageJ software. Multivariate regression models were used to analyze the associations of serum lipid levels with the total HE area and HE location. Results: Sixty-two patients (74 eyes) were enrolled to quantify HEs in DME. The total HE area by MCI was larger than that by CFP (P = 0.004), and the distance between the fovea and the nearest HE by MCI was shorter than that by CFP (P = 0.003). The percentage of patients with HEs involving the central macula by MCI was significantly higher than that by CFP (P < 0.001). Furthermore, 62 eyes of 62 patients were included to analyze the associations of HE parameters with serum lipid levels. In both MCI and CFP, the HE areas were positively associated with triglyceride level (P = 0.016, P = 0.022, respectively). HEs involving the central macula were positively associated with triglyceride and low-density cholesterol levels in MCI (P = 0.028, P = 0.046, respectively), while no significant association was found between serum lipid levels and HE location in CFP. Conclusions: MCI is superior to traditional CFP for the detection of HEs and the analysis of associations between HEs and serum lipid levels in DME. [ABSTRACT FROM AUTHOR]

Introduction: The Ocular manifestations of coronavirus disease 2019 (COVID-19) reported include conjunctivitis, conjunctival hyperemia, chemosis, epiphora, episcleritis, retinal manifestations included cotton wool spots (CWS), micro-hemorrhages, papillophlebitis and neuro-ophthalmic manifestations.Purpose: To report post COVID-19 ophthalmic manifestations using multimodal imaging.Results: A 66-year-old Asian Indian male presented to us with bilateral blurring of vision, RE>LE, of 3 days following a diagnosis of COVID-19 disease. Corrected distance visual acuity were 20/2666 and 20/25 in the right (RE) and left (LE) eyes respectively. He had bilateral anterior chamber inflammation with a relative afferent pupillary defect in the RE. RE showed central retinal artery occlusion(CRAO) with CWS, few flame-shaped retinal hemorrhages and disc edema and hyperemia. LE had disc edema and hyperemia, few flame-shaped retinal hemorrhages, cystoid changes and CWS. A diagnosis of bilateral panuveitis and papillitis with CRAO in the RE was made.Conclusion: Our patient developed a vascular occlusion with panuveitis, which possibly represents an immune mediated event following COVID-19. Patients should be warned about possible ophthalmic sequelae even after recovery. [ABSTRACT FROM AUTHOR]

Purpose: This study was conducted to evaluate the ability of multicolor (MC)–green reflectance (GR) imaging in detecting retinal non-perfused areas (RNPAs) of eyes with diabetic retinopathy (DR). Materials and Methods: Fifty-five degrees fundus fluorescein angiography (FA) and MC imaging were performed in eyes with DR. Images were divided into four fields as macular, nasal, superior temporal, and inferior temporal. To determine the presence and size of RNPAs, FA images were examined by a retina specialist and MC–GR images were examined independently by two masked retina specialists. The compliance of RNPAs in FA and MC–GR images and the agreement of retina specialists were analyzed. Results: The FA and MC–GR images of the 178 eyes were analyzed. When GR images were compared to FA images in terms of RNPAs, sensitivity was 87%; specificity was 89%; the positive predictive value was 92%; the negative predictive value was 83%; and the accuracy was 87%. A substantial agreement was observed between two retina specialists in all retinal fields. Conclusion: This study showed that 55º MC–GR imaging could be an alternative to FA in the evaluation of RNPAs. This study is the first known to explore this issue and needs to be supported and developed with new investigations. [ABSTRACT FROM AUTHOR]

Purpose: To describe multicolor imaging findings in a case of acute retinal pigment epitheliitis. Setting: Vitreoretinal department of a tertiary care center in Eastern India. Methods: A 28-year-old lady presented with acute vision loss in the right eye since 3 days. Left eye examination was unremarkable. Best-corrected visual acuity in the right eye was 20/40. Anterior segment examination revealed clear lens with no evidence of inflammation. There were no vitreous cells. Fundus examination of the right eye revealed a round orange lesion surrounded by a hypopigmented halo in the fovea with multiple tiny hypopigmented lesions temporal to the fovea. Fundus autofluorescence imaging of the right eye showed hyperautofluorescent lesions temporal to the fovea corresponding to the hypopigmented lesions seen temporal to the fovea. Spectral domain optical coherence tomography of the right eye revealed subfoveal hyperreflective lesion in the outer retina with disruption of the ellipsoid zone and interdigitation zone with a linear extension upward. Multicolor composite image of the right eye revealed the foveal lesion as circular red patch with an inferior orange extension. The infrared reflectance image revealed hyperreflective area corresponding to the foveal lesion. The blue and green reflectance images were unremarkable. The patient reported an upper respiratory tract infection 7 days before the visual loss. A diagnosis of acute retinal pigment epitheliitis was made based on history, clinical and imaging findings. Conclusion: This is the first description of multicolor imaging signature of acute retinal pigment epitheliitis lesion in published literature. It seems to be a promising noninvasive imaging modality with a potential to replace color fundus photography. [ABSTRACT FROM AUTHOR]

Purpose: To characterize features of central retinal artery occlusion (CRAO) using multicolor (MC) imaging and to assess the differences in CRAO grading between color fundus photography (CFP) and MC image qualitatively and quantitatively., Methods: We conducted a prospective, cross-sectional study in the Department of Ophthalmology of Renmin Hospital of Wuhan University. In total, 86 acute CRAO patients were included. Spectral-domain optical coherence tomography (SD-OCT), CFP, and MC examinations were taken at baseline. Based on the findings of these three examinations, CRAO was divided into three grades (incomplete, subtotal, and total). Based on OCT grading criteria, we qualitatively compared the ability of grading CRAO by CFP and MC. CRAO patient's visual acuity (VA) was obtained from the initial visit. The retinal thickness was measured by SD-OCT. Superficial capillary plexus (SCP) and deep capillary plexus (DCP) were obtained from optical coherence tomography angiography (OCTA) examinations. Quantitative data were compared across the three acute CRAO subgroups and against three examination findings., Results: MC image had significantly higher power of acute CRAO detection than CFP ( P = 0.03). In the same group of CRAO patients, there was no significant difference in VA when comparing OCT with the MC grading system or with the CFP grading system (all P > 0.05). Significant differences in VA were found between the three CRAO subgroups only under MC grading ( P = 0.016). In incomplete CRAO patients, significant differences were found in central fovea thickness (CFT) when comparing OCT with the CFP grading system ( P = 0.019). In the same group of CRAO patients, there was no significant difference in retinal thickness when comparing OCT with the MC grading system (All P > 0.05). Significance differences in CFT ( P < 0.001), innermost retinal layer (IMRL; P < 0.01), middle retinal layer (MRL; P < 0.001), and outer retinal layer (ORL; P = 0.021) were found between the three CRAO subgroups by MC grading. Vessel density of SCP showed a statistically increased as the severity of three CRAO subgroups ( P = 0.03), whereas DCP did not have significant differences ( P = 0.745). Comparisons were made between the OCT grading method and the MC and CFP grading methods; there is no significant difference in vessel density of SCP and DCP (All P > 0.05)., Conclusion: The images obtained by MC are superior to those obtained by CFP in CRAO grading, retinal thickness, and vessel density measurement. MC imaging may be more capable of CRAO grading than OCT. We recommend MC imaging to determine CRAO severity to guide disease treatment and predict visual prognosis., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Wan, Chen, Li, Yang, Wang, Wang, Li, Yang and Xiao.)

Single-particle tracking (SPT) is a powerful technique to unveil molecular behaviors crucial to the understanding of many biological processes, but it is limited by factors such as probe photostability and spectral orthogonality. To overcome these limitations, we develop upconverting nanoparticles (UCNPs), which are photostable over several hours at the single-particle level, enabling long-term multicolor SPT. We investigate the brightness of core-shell UCNPs as a function of inert shell thickness to minimize particle size while maintaining sufficient signal for SPT. We explore different rare-earth dopants to optimize for the brightest probes and find that UCNPs doped with 2% Tm 3+ /30% Yb 3+ , 10% Er 3+ /90% Yb 3+ , and 15% Tm 3+ /85% Yb 3+ represent the optimal probes for blue, green, and near-infrared emission, respectively. The multiplexed 10 nm probes enable three-color single-particle tracking on live HeLa cells for tens of minutes using a single, near-infrared excitation source. These photostable and multiplexed probes open new avenues for numerous biological applications.

Purpose: To describe various clinical features of idiopathic juxtafoveal retinal telangiectasis group 2A or idiopathic macular telangiectasia type 2 (MacTel) on multicolor imaging (MCI) and compare imaging findings of MacTel on MCI with fundus autofluorescence (FAF). Methods: Patients with a clinical diagnosis of MacTel based on Gass and Blodi's classification were included. FAF and MCI images were graded qualitatively for stage of disease, margins of involvement, hyperautofluorescence on FAF (corresponding retinal atrophy on MCI), and detection of crystals. FAF and MCI were graded quantitatively for the area and number of quadrants involved, hypoautofluorescene on FAF (corresponding intraretinal pigment hyperplasia or retinal pigment epithelium [RPE] atrophy on MCI), and foci of right-angled venules. Results: Seventy-eight eyes of forty five patients were included with both imaging modalities showing no difference with respect to staging of non-proliferative MacTel. Retinal crystals were recognized on MCI but not on FAF. Neurosensory retinal atrophy and subretinal neovascular membranes were detected using MCI with 92.3 and 83.3% sensitivity, respectively. Intraretinal pigmented hyperplasia was more accurately detected (70.1 vs 58.4%) compared with RPE atrophy on MCI. MCI showed larger area of involvement, higher number of quadrants involved (p < 0.001), and better delineation of margins (p = 0.002) compared with FAF. A higher mean number of vessel dipping foci was noted on MCI in comparison with FAF (3.34 vs 3.1). Conclusion: Various parameters were more easily defined using MCI compared with FAF which qualifies MCI as an enface depth-resolved imaging adjunct to conventional multimodal imaging in MacTel. The ability to detect enface as well as cross-sectional imaging features makes MCI a valuable tool in MacTel. [ABSTRACT FROM AUTHOR]

Angioid streaks (AS) are irregular crack-like dehiscences in Bruch’s membrane that are often associated with atrophic degeneration of the overlying retinal pigment epithelium. We herein report multimodal imaging of AS. Multicolor imaging highlighted AS in dark orange color. AS were better visualized in infrared reflectance as compared to green reflectance and blue reflectance. Peau d’orange appearance was seen as alternating dark and bright patches on color fundus photography with corresponding hyporeflective and hyperreflective patches on infrared reflectance. Comet lesions showed increased signal on infrared reflectance and hyperautofluorescence. Multicolor imaging is a non-ive imaging modality which helps in clearly delineating these lesions.

Glaucoma is a progressive optic neuropathy associated with irreversible loss of retinal ganglion cells (RGC). This emblematic localized retinal nerve fiber layer defect (RNFLD) can be the earliest sign to detect the ongoing glaucomatous damage. Slitlamp biomicroscopy [90D & 78D], colour and redfree fundus photography, OCT & HRT are used conventionally for early detection of RNFLD. Multicolour Imaging (MCI) is a new non-invasive retinal imaging modality available in Spectralis platform which simultaneously acquires three reflectance images of the retina using three individual lasers producing a composite image thereby allowing analysis of changes at various levels within the retina. MCI provides sharper image, enables imaging through small pupil and hazy media. Current report describes 2 cases where in MCI proved superior to CFP and conventional redfree photograph in delineating area of RNFLD. This is the first report of MCI in RNFL imaging. The present report highlights the role of MCI in detection of RNFLD.

Interpretation of optic disc cupping and neuroretinal rim is challenging in myopic eyes because of large, shallow, and tilted disc, myopic crescent, and macular degeneration. Color and red-free optic disc photographs do not reveal the cup/disc ratio often in myopic eyes. We report characteristics of multicolor optic disc photography in three myopic eyes. All eyes underwent multicolor imaging Spectralis SDOCT system (Heidelberg Engineering, Germany). Owing to use of three laser lights in a confocal design, multicolor optic disc photography was able to delineate the neuroretinal rim and optic cup more clearly than color and red-free optic disc photographs.

Purpose: The current study compares the ability of multicolor imaging (MCI) to detect the lesions of central serous chorioretinopathy against conventional color fundus photographs (CFP).Methods: It was a retrospective, observational case series of 93 eyes of 58 patients of central serous chorioretinopathy who underwent MCI and CFP. MCI and spectral-domain optical coherence tomography (SD-OCT) were performed using Spectralis SD-OCT system (HRA + OCT). CFP was obtained using FF 450 Plus fundus camera (Carl Zeiss Meditec, Jena, Germany). SD-OCT was considered gold standard for subretinal fluid (SRF) and retinal pigment epithelium detachment (PED). CFP was considered confirmatory investigation for fibrin and blue autofluorescence image (BAF) was considered gold standard to detect retinal pigment epithelium (RPE) atrophy.Results: CFP could detect SRF in 41 (44.1%) eyes. MCI detected SRF in 43 (46.2%) eyes. The sensitivity and specificity of MCI to detect SRF were 70.7% and 94.3%, respectively. PED was detected by CFP in 21 (22.6%) eyes and MCI in 27 (29%) eyes. The sensitivity and specificity of MCI to detect PED were 70% and 97.7% respectively. CFP could pick RPE atrophy in 52 (55.9%) eyes whereas MCI was picked it in 78 (83.9%) of eyes.Conclusion: Both MCI and CFP were inferior to a gold standard in identifying the SRF, PED, and RPE atrophy. However, MCI was better than CFP in comparison with gold standard for these clinical findings in CSC. Thus, MCI seems to be a more valuable imaging tool compared to CFP. [ABSTRACT FROM AUTHOR]

Photobleaching is a major challenge in fluorescence microscopy, in particular if high excitation light intensities are used. Signal‐to‐noise and spatial resolution may be compromised, which limits the amount of information that can be extracted from an image. Photobleaching can be bypassed by using exchangeable labels, which transiently bind to and dissociate from a target, thereby replenishing the destroyed labels with intact ones from a reservoir. Here, we demonstrate confocal and STED microscopy with short, fluorophore‐labeled oligonucleotides that transiently bind to complementary oligonucleotides attached to protein‐specific antibodies. The constant exchange of fluorophore labels in DNA‐based STED imaging bypasses photobleaching that occurs with covalent labels. We show that this concept is suitable for targeted, two‐color STED imaging of whole cells. [ABSTRACT FROM AUTHOR]

Differentiating optic disc edema (ODE) from pseudo optic disc edema (PODE) continues to pose a diagnostic dilemma. Current report highlights the role of multicolor imaging (MC) in differentiating ODE from PODE. Composite multicolor images of the disc in ODE show greenish hyperreflectance that extends beyond the optic disc margins with irregular blurry margins and obscured disc vasculature whereas PODE shows a greenish hyperreflectance with clear and distinct margins and well delineated disc vasculature. MC imaging adds to the present armamentarium of imaging modalities obviating needless neurological evaluation mandatory in a case of true disc edema.

Choroidal nevi are benign fundus lesions that require regular follow with documentation. Conventional color fundus photography (CFP) has traditionally been used to images these lesions. Multicolor imaging (MCI) available on Spectralis spectral domain optical coherence tomography system is increasingly been tested vis-à-vis conventional CFP in various retinal diseases. We present data of the right eye of a 59-year-old gentleman with choroidal nevus who underwent conventional CFP as well as MCI. Nevus appeared orange red on MCI and its size appeared larger than the same measured on conventional CFP. We also report infrared reflectance and near infrared autofluorescence features of choroidal nevus.