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2.
Biomed Opt Express ; 14(9): 4845-4861, 2023 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-37791282

RESUMO

This work compares two tomographic imaging technologies, time-domain full-field optical coherence tomography (FFOCT) working in reflection and optical transmission tomography (OTT), using a new optical setup that combines both. We show that, due to forward-scattering properties, the axial sectioning and contrast in OTT can be optimized by tuning illumination. The influence of sample scattering and thickness are discussed. We illustrate the comparison of the two methods in static (morphology) and dynamic (metabolic contrast) regimes using cell cultures, tissues and entire organisms emphasizing the advantages of both approaches.

3.
Biophys Rep (N Y) ; 3(3): 100119, 2023 Sep 13.
Artigo em Inglês | MEDLINE | ID: mdl-37662577

RESUMO

Viruses have a profound influence on all forms of life, motivating the development of rapid and minimally invasive methods for virus detection. In this study, we present a novel methodology that enables quantitative measurement of the interaction between individual biotic nanoparticles and antibodies in solution. Our approach employs a label-free, full-field common-path interferometric technique to detect and track biotic nanoparticles and their interactions with antibodies. It is based on the interferometric detection of light scattered by viruses in aqueous samples for the detection of individual viruses. We employ single-particle tracking analysis to characterize the size and properties of the detected nanoparticles, and to monitor the changes in their diffusive mobility resulting from interactions. To validate the sensitivity of our detection approach, we distinguish between particles having identical diffusion coefficients but different scattering signals, using DNA-loaded and DNA-devoid capsids of the Escherichia coli T5 virus phage. In addition, we have been able to monitor, in real time, the interaction between the bacteriophage T5 and purified antibodies targeting its major capsid protein pb8, as well as between the phage SPP1 and nonpurified anti-SPP1 antibodies present in rabbit serum. Interestingly, these virus-antibody interactions are observed within minutes. Finally, by estimating the number of viral particles interacting with antibodies at different concentrations, we successfully quantify the dissociation constant Kd of the virus-antibody reaction using single-particle tracking analysis.

4.
Front Cell Infect Microbiol ; 13: 1183340, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37502605

RESUMO

Introduction: The diagnosis of cutaneous manifestations of deep mycoses relies on both histopathological and direct examinations. Yet, the current diagnostic criteria cannot prevent missed cases, including invasive aspergillosis, which requires the development of a novel diagnostic approach and imaging tools. We recently introduced the use of dynamic full-field optical coherence tomography (D-FF-OCT) in fungal diagnostics with a definition approaching that of conventional microscopy and the ability to return metabolic information regarding different fungal species. The present work focuses on subcellular dynamics and live-cell imaging of Aspergillus fumigatus with D-FF-OCT to follow the fungal growth stages. Methods: The A. fumigatus ATCC 204305 quality-control strain was used for all imaging experiments, following incubation times varying between 24 and 72 h at 30°C in a humidified chamber on Sabouraud dextrose agar. Fungal growth was subsequently monitored with D-FF-OCT for up to 5 h at room temperature and following the pharmacological stress of either voriconazole, amphotericin B, or caspofungin gradient concentration. Results: D-FF-OCT images allow not only the visualization of intracellular trafficking of vacuoles but also an evolving dynamic segmentation of conidiophores depending on the chronological development and aging of the hyphae or the effect of antifungal treatment. The same applies to conidial heads, with the most intense D-FF-OCT signal coming from vesicles, revealing a changing dynamic within a few hours only, as well as complete extinction following subsequent drying of the Sabouraud dextrose agar. Discussion: These results provide additional data on the ability of D-FF-OCT to monitor some of the main life cycle processes, dynamics, and intracellular trafficking of vacuoles in A. fumigatus, with or without the effect of pharmacological stress. Such complementary metabolic information could help both clinicians and microbiologists in either mechanistic studies toward experimental mycology or the development of a potential D-FF-OCT-guided diagnosis of superficial fungal infections.


Assuntos
Aspergillus fumigatus , Tomografia de Coerência Óptica , Ágar/farmacologia , Antifúngicos/farmacologia , Glucose
5.
J Med Imaging (Bellingham) ; 10(3): 034504, 2023 May.
Artigo em Inglês | MEDLINE | ID: mdl-37274760

RESUMO

Purpose: The adoption of emerging imaging technologies in the medical community is often hampered when they provide a new unfamiliar contrast that requires experience to be interpreted. Dynamic full-field optical coherence tomography (D-FF-OCT) microscopy is such an emerging technique. It provides fast, high-resolution images of excised tissues with a contrast comparable to H&E histology but without any tissue preparation and alteration. Approach: We designed and compared two machine learning approaches to support interpretation of D-FF-OCT images of breast surgical specimens and thus provide tools to facilitate medical adoption. We conducted a pilot study on 51 breast lumpectomy and mastectomy surgical specimens and more than 1000 individual 1.3×1.3 mm2 images and compared with standard H&E histology diagnosis. Results: Using our automatic diagnosis algorithms, we obtained an accuracy above 88% at the image level (1.3×1.3 mm2) and above 96% at the specimen level (above cm2). Conclusions: Altogether, these results demonstrate the high potential of D-FF-OCT coupled to machine learning to provide a rapid, automatic, and accurate histopathology diagnosis with minimal sample alteration.

7.
Biomed Opt Express ; 13(8): 4190-4203, 2022 Aug 01.
Artigo em Inglês | MEDLINE | ID: mdl-36032580

RESUMO

There is an increasing need for label free methods that could reveal intracellular structures and dynamics. In this context, we develop a new optical tomography method working in transmission - full-field optical transmission tomography (FF-OTT). The method can measure the forward scattering signals and reveals the time-dependent metabolic signals in living cells. FF-OTT is a common path interferometer taking advantage of the Gouy phase shift - a π phase shift that the light wave experiences around the focus. By modulating the position of the focus one can alter the phase of the scattered light. Demodulation of images with different phases rejects the background and enhances the light from the depth-of-field, thus producing an optical section. We test FF-OTT by imaging single-cell diatoms and ex vivo biological samples. In fresh samples, we show that the intracellular motions create visible intensity fluctuations in FF-OTT so that the method is able to reveal a metabolic dynamic contrast. FF-OTT was found to be an efficient label free technique that can be readily implemented thanks to a robust common-path speckle-free interferometer design using an incoherent light source.

8.
Biomed Opt Express ; 13(6): 3187-3194, 2022 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-35781969

RESUMO

We present a novel method that can assay cellular viability in real-time using supervised machine learning and intracellular dynamic activity data that is acquired in a label-free, non-invasive, and non-destructive manner. Cell viability can be an indicator for cytology, treatment, and diagnosis of diseases. We applied four supervised machine learning models on the observed data and compared the results with a trypan blue assay. The cell death assay performance by the four supervised models had a balanced accuracy of 93.92 ± 0.86%. Unlike staining techniques, where criteria for determining viability of cells is unclear, cell viability assessment using machine learning could be clearly quantified.

9.
Commun Biol ; 5(1): 575, 2022 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-35688936

RESUMO

Retinal degenerative diseases lead to the blindness of millions of people around the world. In case of age-related macular degeneration (AMD), the atrophy of retinal pigment epithelium (RPE) precedes neural dystrophy. But as crucial as understanding both healthy and pathological RPE cell physiology is for those diseases, no current technique allows subcellular in vivo or in vitro live observation of this critical cell layer. To fill this gap, we propose dynamic full-field OCT (D-FFOCT) as a candidate for live observation of in vitro RPE phenotype. In this way, we monitored primary porcine and human stem cell-derived RPE cells in stress model conditions by performing scratch assays. In this study, we quantified wound healing parameters on the stressed RPE, and observed different cell phenotypes, displayed by the D-FFOCT signal. In order to decipher the subcellular contributions to these dynamic profiles, we performed immunohistochemistry to identify which organelles generate the signal and found mitochondria to be the main contributor to D-FFOCT contrast. Altogether, D-FFOCT appears to be an innovative method to follow degenerative disease evolution and could be an appreciated method in the future for live patient diagnostics and to direct treatment choice.


Assuntos
Degeneração Macular , Epitélio Pigmentado da Retina , Animais , Angiofluoresceinografia , Humanos , Degeneração Macular/metabolismo , Mitocôndrias , Epitélio Pigmentado da Retina/metabolismo , Suínos , Tomografia de Coerência Óptica/métodos
10.
J Mycol Med ; 32(4): 101303, 2022 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-35732095

RESUMO

Histopathology and microscopic examination of infected tissue are the gold standards to prove the diagnosis of invasive fungal infection (IFI). Yet, they suffer from essential limitations that hamper rapid diagnosis and require the future development of new imaging tools dedicated to fungal diagnostics. To this end, the present work introduces the first use of dynamic full-field optical coherence tomography (D-FF-OCT) for the visualization of microscopic filamentous fungi. Data collected from the observation of three different fungal species (Nannizzia gypsea, Aspergillus fumigatus and Rhizopus arrhizus) confirm the ability of D-FF-OCT to visualize not only the main structures of all selected fungal species (hyphae, spores, conidia, sporulating structures), but also the metabolic activity of the organisms, which could provide additional help in the future to better characterize the signature of each fungal structure. These results demonstrate how D-FF-OCT could serve as potential complementary tool for rapid diagnosis of IFI in both intensive and non-intensive care units.


Assuntos
Aspergillus fumigatus , Tomografia de Coerência Óptica , Tomografia de Coerência Óptica/métodos , Esporos Fúngicos , Fungos
11.
Biomed Opt Express ; 12(10): 6431-6441, 2021 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-34745747

RESUMO

Dynamic full-field optical coherence microscopy (DFFOCM) was used to characterize the intracellular dynamic activities and cytoskeleton of HeLa cells in different viability states. HeLa cell samples were continuously monitored for 24 hours and compared with histological examination to confirm the cell viability states. The averaged mean frequency and magnitude observed in healthy cells were 4.79±0.5 Hz and 2.44±1.06, respectively. In dead cells, the averaged mean frequency was shifted to 8.57±0.71 Hz, whereas the magnitude was significantly decreased to 0.53±0.25. This cell dynamic activity analysis using DFFOCM is expected to replace conventional time-consuming and biopsies-required histological or biochemical methods.

12.
Gut ; 70(1): 6-8, 2021 01.
Artigo em Inglês | MEDLINE | ID: mdl-32447309

RESUMO

Full-field optical coherence tomography (FFOCT) is an imaging technique of biological tissue based on tissue light reflectance analysis. We evaluated the feasibility of imaging fresh digestive mucosal biopsies after a quick mounting procedure (5 min) using two distinct modalities of FFOCT. In static FFOCT mode, we gained high-resolution images of general gut tissue-specific architecture, such as oesophageal papillae, gastric pits, duodenal villi and colonic crypts. In dynamic FFOCT mode, we imaged individual epithelial cells of the mucosal lining with a cellular or subcellular resolution and identified cellular components of the lamina propria. FFOCT represents a promising dye-free imaging tool for on-site analysis of gut tissue remodelling.


Assuntos
Trato Gastrointestinal/diagnóstico por imagem , Trato Gastrointestinal/patologia , Tomografia de Coerência Óptica , Biópsia , Endoscopia , Humanos , Mucosa/diagnóstico por imagem , Mucosa/patologia
13.
Opt Lett ; 45(21): 5901-5904, 2020 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-33137028

RESUMO

The highest three-dimensional (3D) resolution possible in in vivo retinal imaging is achieved by combining optical coherence tomography (OCT) and adaptive optics. However, this combination brings important limitations, such as small field-of-view and complex, cumbersome systems, preventing so far the translation of this technology from the research lab to clinics. In this Letter, we mitigate these limitations by combining our compact time-domain full-field OCT (FFOCT) with a multi-actuator adaptive lens positioned just in front of the eye, in a technique we call the adaptive-glasses wavefront sensorless approach. Through this approach, we demonstrate that ocular aberrations can be corrected, increasing the FFOCT signal-to-noise ratio (SNR) and enabling imaging of different retinal layers with a 3D cellular resolution over a 5∘×5∘ field-of-view, without apparent anisoplanatism.


Assuntos
Lentes , Retina/diagnóstico por imagem , Razão Sinal-Ruído , Tomografia de Coerência Óptica/instrumentação , Humanos , Fatores de Tempo
14.
Biomed Opt Express ; 11(9): 4928-4941, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-33014591

RESUMO

Allying high-resolution with a large field-of-view (FOV) is of great importance in the fields of biology and medicine, but it is particularly challenging when imaging non-flat living samples such as the human retina. Indeed, high-resolution is normally achieved with adaptive optics (AO) and scanning methods, which considerably reduce the useful FOV and increase the system complexity. An alternative technique is time-domain full-field optical coherence tomography (FF-OCT), which has already shown its potential for in-vivo high-resolution retinal imaging. Here, we introduce coherence gate shaping for FF-OCT, to optically shape the coherence gate geometry to match the sample curvature, thus achieving a larger FOV than previously possible. Using this instrument, we obtained high-resolution images of living human photoreceptors close to the foveal center without AO and with a 1 mm × 1 mm FOV in a single shot. This novel advance enables the extraction of photoreceptor-based biomarkers with ease and spatiotemporal monitoring of individual photoreceptors. We compare our findings with AO-assisted ophthalmoscopes, highlighting the potential of FF-OCT, as a compact system, to become a routine clinical imaging technique.

15.
Light Sci Appl ; 9: 140, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32864115

RESUMO

Optical coherence tomography offers astounding opportunities to image the complex structure of living tissue but lacks functional information. We present dynamic full-field optical coherence tomography as a technique to noninvasively image living human induced pluripotent stem cell-derived retinal organoids. Coloured images with an endogenous contrast linked to organelle motility are generated, with submicrometre spatial resolution and millisecond temporal resolution, creating a way to identify specific cell types in living tissue via their function.

16.
PLoS One ; 15(8): e0234165, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32866179

RESUMO

Histopathological examination of temporal artery biopsy (TAB) remains the gold standard for the diagnosis of giant cell arteritis (GCA) but is associated with essential limitations that emphasize the need for an upgraded pathological process. This study pioneered the use of full-field optical coherence tomography (FF-OCT) for rapid and automated on-site pathological diagnosis of GCA. Sixteen TABs (12 negative and 4 positive for GCA) were selected according to major histopathological criteria of GCA following hematoxylin-eosin-saffron-staining for subsequent acquisition with FF-OCT to compare structural modifications of the artery cell wall and thickness of each tunica. Gabor filtering of FF-OCT images was then used to compute TAB orientation maps and validate a potential automated analysis of TAB sections. FF-OCT allowed both qualitative and quantitative visualization of the main structures of the temporal artery wall, from the internal elastic lamina to the vasa vasorum and red blood cells, unveiling a significant correlation with conventional histology. FF-OCT imaging of GCA TABs revealed destruction of the media with distinct remodeling of the whole arterial wall into a denser reticular fibrous neo-intima, which is distinctive of GCA pathogenesis and accessible through automated Gabor filtering. Rapid on-site FF-OCT TAB acquisition makes it possible to identify some characteristic pathological lesions of GCA within a few minutes, paving the way for potential machine intelligence-based or even non-invasive diagnosis of GCA.


Assuntos
Arterite de Células Gigantes/diagnóstico por imagem , Tomografia de Coerência Óptica/métodos , Idoso , Idoso de 80 Anos ou mais , Estudos de Casos e Controles , Estudos de Coortes , Feminino , Arterite de Células Gigantes/diagnóstico , Arterite de Células Gigantes/patologia , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Prospectivos , Artérias Temporais/diagnóstico por imagem , Artérias Temporais/patologia , Túnica Íntima/diagnóstico por imagem , Túnica Íntima/patologia , Túnica Média/diagnóstico por imagem , Túnica Média/patologia
17.
Biomed Opt Express ; 11(1): 492-504, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-32010530

RESUMO

Time-domain full-field OCT (FF-OCT) represents an imaging modality capable of recording high-speed en-face sections of a sample at a given depth. One of the biggest challenges to transfer this technique to image in-vivo human retina is the presence of continuous involuntary head and eye axial motion during image acquisition. In this paper, we demonstrate a solution to this problem by implementing an optical stabilization in an FF-OCT system. This was made possible by combining an FF-OCT system, an SD-OCT system, and a high-speed voice-coil translation stage. B-scans generated by the SD-OCT were used to measure the retina axial position and to drive the position of the high-speed voice coil translation stage, where the FF-OCT reference arm is mounted. Closed-loop optical stabilization reduced the RMS error by a factor of 7, significantly increasing the FF-OCT image acquisition efficiency. By these means, we demonstrate the capacity of the FF-OCT to resolve cone mosaic as close as 1.5 o from the fovea center with high consistency and without using adaptive optics.

18.
Biomed Opt Express ; 10(2): 731-746, 2019 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-30800511

RESUMO

We describe recent technological progress in multimodal en face full-field optical coherence tomography that has allowed detection of slow and fast dynamic processes in the eye. We show that by combining static, dynamic and fluorescence contrasts we can achieve label-free high-resolution imaging of the retina and anterior eye with temporal resolution from milliseconds to several hours, allowing us to probe biological activity at subcellular scales inside 3D bulk tissue. Our setups combine high lateral resolution over a large field of view with acquisition at several hundreds of frames per second which make it a promising tool for clinical applications and biomedical studies. Its contactless and non-destructive nature is shown to be effective for both following in vitro sample evolution over long periods of time and for imaging of the human eye in vivo.

19.
Nat Commun ; 9(1): 3804, 2018 09 18.
Artigo em Inglês | MEDLINE | ID: mdl-30228263

RESUMO

Defects in cerebrospinal fluid (CSF) flow may contribute to idiopathic scoliosis. However, the mechanisms underlying detection of CSF flow in the central canal of the spinal cord are unknown. Here we demonstrate that CSF flows bidirectionally along the antero-posterior axis in the central canal of zebrafish embryos. In the cfap298tm304 mutant, reduction of cilia motility slows transport posteriorly down the central canal and abolishes spontaneous activity of CSF-contacting neurons (CSF-cNs). Loss of the sensory Pkd2l1 channel nearly abolishes CSF-cN calcium activity and single channel opening. Recording from isolated CSF-cNs in vitro, we show that CSF-cNs are mechanosensory and require Pkd2l1 to respond to pressure. Additionally, adult pkd2l1 mutant zebrafish develop an exaggerated spine curvature, reminiscent of kyphosis in humans. These results indicate that CSF-cNs are mechanosensory cells whose Pkd2l1-driven spontaneous activity reflects CSF flow in vivo. Furthermore, Pkd2l1 in CSF-cNs contributes to maintenance of natural curvature of the spine.


Assuntos
Líquido Cefalorraquidiano/metabolismo , Mecanotransdução Celular , Neurônios/metabolismo , Medula Espinal/metabolismo , Proteínas de Peixe-Zebra/metabolismo , Peixe-Zebra/metabolismo , Animais , Cílios/metabolismo
20.
J Cancer Res Clin Oncol ; 144(10): 1967-1990, 2018 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-29926160

RESUMO

INTRODUCTION: Tumor detection and visualization plays a key role in the clinical workflow of a patient with suspected cancer, both in the diagnosis and treatment. Several optical imaging techniques have been evaluated for guidance during oncological interventions. Optical coherence tomography (OCT) is a technique which has been widely evaluated during the past decades. This review aims to determine the clinical usefulness of OCT during cancer interventions focussing on qualitative features, quantitative features and the diagnostic value of OCT. METHODS: A systematic literature search was performed for articles published before May 2018 using OCT in the field of surgical oncology. Based on these articles, an overview of the clinical usefulness of OCT was provided per tumor type. RESULTS: A total of 785 articles were revealed by our search, of which a total of 136 original articles were available for analysis, which formed the basis of this review. OCT is currently utilised for both preoperative diagnosis and intraoperative detection of skin, oral, lung, breast, hepatobiliary, gastrointestinal, urological, and gynaecological malignancies. It showed promising results in tumor detection on a microscopic level, especially using higher resolution imaging techniques, such as high-definition OCT and full-field OCT. CONCLUSION: In the near future, OCT could be used as an additional tool during bronchoscopic or endoscopic interventions and could also be implemented in margin assessment during (laparoscopic) cancer surgery if a laparoscopic or handheld OCT device will be further developed to make routine clinical use possible.


Assuntos
Neoplasias/diagnóstico por imagem , Neoplasias/cirurgia , Cirurgia Assistida por Computador/métodos , Tomografia de Coerência Óptica/métodos , Humanos , Oncologia Cirúrgica/métodos
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