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1.
J Biomed Opt ; 29(Suppl 2): S22702, 2025 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-38434231

RESUMO

Significance: Advancements in label-free microscopy could provide real-time, non-invasive imaging with unique sources of contrast and automated standardized analysis to characterize heterogeneous and dynamic biological processes. These tools would overcome challenges with widely used methods that are destructive (e.g., histology, flow cytometry) or lack cellular resolution (e.g., plate-based assays, whole animal bioluminescence imaging). Aim: This perspective aims to (1) justify the need for label-free microscopy to track heterogeneous cellular functions over time and space within unperturbed systems and (2) recommend improvements regarding instrumentation, image analysis, and image interpretation to address these needs. Approach: Three key research areas (cancer research, autoimmune disease, and tissue and cell engineering) are considered to support the need for label-free microscopy to characterize heterogeneity and dynamics within biological systems. Based on the strengths (e.g., multiple sources of molecular contrast, non-invasive monitoring) and weaknesses (e.g., imaging depth, image interpretation) of several label-free microscopy modalities, improvements for future imaging systems are recommended. Conclusion: Improvements in instrumentation including strategies that increase resolution and imaging speed, standardization and centralization of image analysis tools, and robust data validation and interpretation will expand the applications of label-free microscopy to study heterogeneous and dynamic biological systems.


Assuntos
Técnicas Histológicas , Microscopia , Animais , Citometria de Fluxo , Processamento de Imagem Assistida por Computador
2.
Acta Neurochir (Wien) ; 166(1): 118, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38427127

RESUMO

BACKGROUND: The surgical 3D exoscopes have recently been introduced as an alternative to the surgical microscopes in microneurosurgery. Since the exoscope availability is still limited, it is relevant to know whether even a short-term exoscope training develops the skills needed for performing exoscope-assisted surgeries. METHODS: Ten participants (six consultants, four residents) performed two laboratory bypass test tasks with a 3D exoscope (Aesculap Aeos®). Six training sessions (6 h) were performed in between (interval of 2-5 weeks) on artificial models. The participants were divided into two groups: test group (n = 6) trained with the exoscope and control group (n = 4) with a surgical microscope. The test task was an artificial end-to-side microsurgical anastomosis model, using 12 interrupted 9-0 sutures and recorded on video. We compared the individual as well as group performance among the test subjects based on suturing time, anastomosis quality, and manual dexterity. RESULTS: Altogether, 20 bypass tasks were performed (baseline n = 10, follow-up n = 10). The median duration decreased by 28 min and 44% in the exoscope training group. The decrease was steeper (29 min, 45%) among the participants with less than 6 years of microneurosurgery experience compared to the more experienced participants (13 min, 24%). After training, the participants with at least 1-year experience of using the exoscope did not improve their task duration. The training with the exoscope led to a greater time reduction than the training with the microscope (44% vs 17%). CONCLUSIONS: Even short-term training with the exoscope led to marked improvements in exoscope-assisted bypass suturing among novice microneurosurgeons. For the more experienced participants, a plateau in the initial learning curve was reached quickly. A much longer-term effort might be needed to witness further improvement in this user group.


Assuntos
Microcirurgia , Procedimentos Neurocirúrgicos , Humanos , Estudos Prospectivos , Microscopia
3.
Methods Mol Biol ; 2761: 589-597, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38427263

RESUMO

Immunolabeling-enabled imaging of solvent-cleared organs (iDISCO) (Renier N, Wu Z, Simon DJ, Yang J, Ariel P, Tessier-Lavigne M, Cell 159:896-910, 2014) aims to match the refractive index (RI) of tissue to the surrounding medium, thereby facilitating three-dimensional (3D) imaging and quantification of cellular points and tissue structures. Once cleared, transparent tissue samples allow for rapid imaging with no mechanical sectioning. This imaging technology enables us to visualize brain tissue in situ and quantify the morphology and extent of glial cell branches or neuronal processes extending from the epicenter of a traumatic brain injury (TBI). In this way, we can more accurately assess and quantify the damaging consequences of TBI not only in the impact region but also in the extended pericontusional regions.


Assuntos
Lesões Encefálicas Traumáticas , Microscopia , Camundongos , Animais , Imageamento Tridimensional/métodos , Solventes , Lesões Encefálicas Traumáticas/diagnóstico por imagem , Encéfalo
4.
Development ; 151(5)2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38477686

RESUMO

Observation is the heart of research, but it can be challenging to observe deeply and go beyond expected observations. Here, we describe activities designed for scientists to enhance their observational skills by engaging with art. In collaboration with an art gallery at our university, our lab practiced observing representational paintings in a systematic way, separating the act of observation from interpretation. Applying this skill to our microscopy images allowed us to access information in the data that may otherwise have been overlooked. In addition, these activities highlighted the power of collecting observations from multiple observers before generating interpretations, as well as the value of discussing the creative and emotional aspects of data collection and interpretation. We provide concrete examples of how we will incorporate these skills into our research processes, as well as details that other groups can use to engage in similar art-based training activities to enhance their own observational skills.


Assuntos
Coração , Microscopia , Humanos
5.
Brief Bioinform ; 25(2)2024 Jan 22.
Artigo em Inglês | MEDLINE | ID: mdl-38483256

RESUMO

Numerous imaging techniques are available for observing and interrogating biological samples, and several of them can be used consecutively to enable correlative analysis of different image modalities with varying resolutions and the inclusion of structural or molecular information. Achieving accurate registration of multimodal images is essential for the correlative analysis process, but it remains a challenging computer vision task with no widely accepted solution. Moreover, supervised registration methods require annotated data produced by experts, which is limited. To address this challenge, we propose a general unsupervised pipeline for multimodal image registration using deep learning. We provide a comprehensive evaluation of the proposed pipeline versus the current state-of-the-art image registration and style transfer methods on four types of biological problems utilizing different microscopy modalities. We found that style transfer of modality domains paired with fully unsupervised training leads to comparable image registration accuracy to supervised methods and, most importantly, does not require human intervention.


Assuntos
Aprendizado Profundo , Humanos , Microscopia
6.
Zootaxa ; 5415(1): 144-152, 2024 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-38480211

RESUMO

Diadelophora gen. nov, a conspicuous new genus of phorid flies is described based on two species from central and western Brazil, D. stilbella sp. nov. and D. inornata sp. nov. The new genus is positioned in the Thaumatoxena-group within the subfamily Phorinae, probably as sister group to Hypocerides Schmitz, 1915. The diagnostic features of Diadelophora are commented and illustrated, and the genus differences to Hypocerides are highlighted. The morphology of Diadelophora species is explored in detail with SEM, photos, and optical microscopy illustrations of structures of taxonomic relevance and other curious features of the genus.


Assuntos
Dípteros , Animais , Brasil , Microscopia , Distribuição Animal
7.
Zootaxa ; 5408(1): 1-184, 2024 Feb 09.
Artigo em Inglês | MEDLINE | ID: mdl-38480261

RESUMO

The Australian Lasioglossum Curtis 1833 subgenus Parasphecodes Smith 1853 is revised. Currently, Parasphecodes has 92 named, described species. The monotypic Lasioglossum subgenus Pseudochilalictus Michener 1965 is synonymised with Parasphecodes and its species, L. imitator Michener 1965, is recombined into Parasphecodes. The single known species from New Guinea, L. (Parasphecodes) permetallicum Michener 1965, is included in this revision. Eighteen new species are erected, 69 names are placed into synonymy, 20 new sex associations made and three species currently placed in Parasphecodes are recombined into the Lasioglossum subgenus Ctenonomia Cameron 1903. This revision resolved there are 40 valid species for Parasphecodes. Valid species for Lasioglossum (Parasphecodes) without synonymies are as follows: L. imitator, L. lichatus (Smith 1853), L. loweri (Cockerell 1905), L. olgae (Rayment 1935), L. permetallicum, L. turneri (Cockerell 1914d) and L. waterhousei (Cockerell 1915a). New synonymies proposed for Lasioglossum (Parasphecodes) are as follows: Lasioglossum (Pseudochilalictus Michener 1965) new synonymy = Lasioglossum (Parasphecodes); L. cirriferum (Cockerell 1910) new synonymy, L. insigne (Meyer 1920) new synonymy and L. grande (Meyer 1920) new synonymy = L. altichus (Smith 1853); L. paramelaenum (Cockerell 1922) new synonymy = L. atronitens (Cockerell 1914a); L. bribiense (Cockerell 1916) new synonymy, L. bribiensiforme (Cockerell 1930a) new synonymy, L. butleri (Rayment 1935) new synonymy, L. frenchi (Rayment 1935) new synonymy, L. frenchellum Michener 1965 new synonymy, L. sordidulum (Cockerell 1914c) new synonymy and L. patongensis (Rayment 1948) new synonymy = L. bryotrichum (Cockerell 1912a); L. fumidicaudum (Cockerell 1914b) new synonymy and L. noachinum (Cockerell 1914b) new synonymy = L. carbonarium (Smith 1853); L. cervicale (Cockerell 1915b) new synonymy and L. zamelanum (Cockerell 1930a) new synonymy = L. dissimulator (Cockerell 1914b); L. wilmatae (Cockerell 1929c) new synonymy = L. excultum (Cockerell 1913b); L. arciferum (Cockerell 1914b) new synonymy, L. atrorufescens (Cockerell 1914b) new synonymy, L. fulviventre (Friese 1924) new synonymy, L. leptospermi (Cockerell 1916) new synonymy, L. lichatinum (Cockerell 1922) new synonymy, L. leucorhinum (Cockerell 1926) new synonymy, L. proximum (Rayment 1947) new synonymy, L. testaciventre (Rayment 1953) new synonymy, L. tilachus (Smith 1853) new synonymy, L. tilachiforme (Cockerell 1907) new synonymy, L. tuchilas (Smith 1853) new synonymy, L. anhybodinum (Cockerell 1930a) new synonymy, L. hybodinum (Cockerell 1912a) new synonymy, L. tripunctatum (Cockerell 1929c) new synonymy and L. warburtoni (Cockerell 1906) new synonymy = L. hilactus (Smith 1853); L. frenchi (Cockerell 1904) new synonymy, L. schomburgki (Cockerell 1910) new synonymy, L. speculiferum (Cockerell 1912a) new synonymy, L. sextum (Cockerell 1910) new synonymy, L. solis (Cockerell 1922) new synonymy, L. vermiculatum (Cockerell 1914b) new synonymy and L. vulneratum (Cockerell 1910) new synonymy = L. hiltacus (Smith 1853); L. hirtiventre (Cockerell 1922) new synonymy, L. niveorufum (Friese 1924) new synonymy, L. submeracum (Cockerell 1930a) new synonymy and L. froggatti (Cockerell 1905) new synonymy = L. lacthius (Smith 1853); L. basilautum (Cockerell 1910) new synonymy, L. doddi (Cockerell 1914c) new synonymy, L. paracolletinum (Cockerell 1910) new synonymy, L. pilicolle (Friese 1924) new synonymy, L. scutellatum (Friese 1924) new synonymy and L. vau (Cockerell 1910) new synonymy = L. leichardti (Cockerell 1906); L. annexum (Cockerell 1922) new synonymy, L. latissimum (Cockerell 1915b) new synonymy, L. microdontum (Cockerell 1912a) new synonymy, L. recessum (Cockerell 1914d) new synonymy, L. longmani (Cockerell 1922) new synonymy, L. recantans (Cockerell 1912a) new synonymy and L. rufotegulare (Cockerell 1914e) new synonymy = L. melbournense (Cockerell 1904); L. trimaculatum (Friese 1924) new synonymy = L. musicum (Cockerell 1913a); L. gentianae (Rayment 1951) new synonymy = L. subrussatum (Cockerell 1922); L. fultoni (Cockerell 1914b) new synonymy, L. gibbosum (Friese 1924) new synonymy, L. niveatum (Meyer 1920) new synonymy, L. punctatissimus (Meyer 1920) new synonymy, L. rhodopterum (Cockerell 1914e) new synonymy, L. rubriventre (Friese 1924) new synonymy, L. subfultoni (Cockerell 1930a) new synonymy, L. tepperi (Cockerell 1905) new synonymy, L. notescens (Cockerell 1930a) new synonymy and L. rufulum (Friese 1924) new synonymy = L. sulthica (Smith 1853); L. submoratum (Cockerell 1930a) new synonymy and L. perustum (Cockerell 1914d) new synonymy = L. taluchis (Smith 1853). Eighteen new species are described as follows: L. acristum Walker & Sparks, L. altum Walker & Sparks, L. aspereticulum Walker & Sparks, L. atropum Walker & Sparks, L. bimelasmum Walker & Sparks, L. bipenicillum Walker & Sparks, L. bitrichum Walker & Sparks, L. blyscanatum Walker & Sparks, L. brevipectinatum Walker & Sparks, L. capronum Walker & Sparks, L. ferruginum Walker & Sparks, L. flexosum Walker & Sparks, L. laevidiscum Walker & Sparks, L. recavum Walker & Sparks, L. reticulum Walker & Sparks, L. rutrum Walker & Sparks, L. variegatum Walker & Sparks and L. wcisloi Walker & Sparks. New subgeneric classifications are as follows: L. (Pseudochilalictus) imitator = L. (Parasphecodes) imitator new status, Halictus clarigaster Cockerell 1918 = L. (Ctenonomia) clarigaster new status, Halictus forresti Cockerell 1906 = L. (Ctenonomia) forresti new status, and Halictus tribuarius Rayment 1935 = L. (Ctenonomia) tribuarium new status. These species names, all described by Smith 1853, are anagrams of Halictus. Therefore, they are nouns in apposition and should retain their original species designations as: Lasioglossum (Parasphecodes) altichus (Smith 1853), Lasioglossum (Parasphecodes) hilactus (Smith 1853), Lasioglossum (Parasphecodes) hiltacus (Smith 1853), Lasioglossum (Parasphecodes) lacthius (Smith 1853), Lasioglossum (Parasphecodes) lichatus (Smith 1853), Lasioglossum (Parasphecodes) sulthica (Smith 1853), Lasioglossum (Parasphecodes) talchius (Smith 1853), Lasioglossum (Parasphecodes) taluchis (Smith 1853), Lasioglossum (Parasphecodes) tilachus (Smith 1853) and Lasioglossum (Parasphecodes) tuchilas (Smith 1853). All 40 valid Parasphecodes species, as well as the three species recombined to Ctenonomia, are redescribed. For the Parasphecodes species, keys to both sexes, character groups, taxonomy, citations, species diagnoses, comments, descriptions, scanning electron micrographs, colour montage images, distribution maps, male genitalia and S7S8 line drawings are provided to assist with species identifications.


Assuntos
Himenópteros , Feminino , Abelhas , Masculino , Animais , Austrália , Distribuição Animal , Microscopia
8.
Zootaxa ; 5405(1): 142-150, 2024 Jan 26.
Artigo em Inglês | MEDLINE | ID: mdl-38480392

RESUMO

Raphidocystis marginata (Siemensma, 1981), Raineriophrys echinata (Rainer, 1968), and Pterocystis fortesca (Nicholls, 1983) are heliozoan protists, have been recorded only in a few localities in Europe, and considered to be rare species. These centrohelid heliozoans have been reported for the first time in Ukrainian Polissia, and we provide their morphological descriptions with new morphometric data of exoskeleton (periplast) based on Ukrainian material. The diagnostic morphological characters are illustrated by light and scanning electron microscope photographs. Their geographical distribution in Europe and biotope preference are discussed.


Assuntos
Eucariotos , Microscopia , Animais , Europa (Continente)
9.
J Biomed Opt ; 29(Suppl 1): S11527, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38464883

RESUMO

Significance: We developed a high-speed optical-resolution photoacoustic microscopy (OR-PAM) system using a high-repetition-rate supercontinuum (SC) light source and a two-axes Galvano scanner. The OR-PAM system enabled real-time imaging of optical absorbers inside biological tissues with excellent excitation wavelength tunability. Aim: In the near-infrared (NIR) wavelength range, high-speed OR-PAM faces limitations due to the lack of wavelength-tunable light sources. Our study aimed to enable high-speed OR-PAM imaging of various optical absorbers, including NIR contrast agents, and validate the performance of high-speed OR-PAM in the detection of circulating tumor cells (CTCs). Approach: A high-repetition nanosecond pulsed SC light source was used for OR-PAM. The excitation wavelength was adjusted by bandpass filtering of broadband light pulses produced by an SC light source. Phantom and in vivo experiments were performed to detect tumor cells stained with an NIR contrast agent within flowing blood samples. Results: The newly developed high-speed OR-PAM successfully detected stained cells both in the phantom and in vivo. The phantom experiment confirmed the correlation between the tumor cell detection rate and tumor cell concentration in the blood sample. Conclusions: The high-speed OR-PAM effectively detected stained tumor cells. Combining high-speed OR-PAM with molecular probes that stain tumor cells in vivo enables in vivo CTC detection.


Assuntos
Dispositivos Ópticos , Técnicas Fotoacústicas , Microscopia/métodos , Técnicas Fotoacústicas/métodos , Análise Espectral , Imagens de Fantasmas
10.
Anal Chim Acta ; 1297: 342345, 2024 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-38438227

RESUMO

Mass spectrometry imaging (MSI) visualizes spatial distribution of molecules in a biological tissue. However, compared with traditional microscopy-based imaging, conventional MSI is limited to its spatial resolution, resulting in difficulties in identifying detailed tissue morphological characters, such as lesion boundary or nanoscale structures. On the other hand, expansion microscopy, a tissue expansion method widely used in optical imaging to improve morphological details, has great potential to solve insufficient spatial resolution in mass spectrometry imaging (MSI). However, expansion microscopy was not originally designed for MSI, resulting in problems while combining expansion microscopy and MSI such as expanded sample fragility, vacuum stability and molecule loss during sample preparation. In this research we developed a MALDI MSI compatible expansion protocol by adjusting sample preparation methods during tissue expansion, successfully combining expansion microscopy with MSI. After tissue expansion the expanded sample can be readily applied to MALDI MSI sample preparation and further data acquisition. The MALDI MSI compatible expansion protocol has great potential to be widely applied in MALDI MSI sample preparation to facilitate improvement of MSI spatial resolution.


Assuntos
Microscopia , Imagem Óptica , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Expansão de Tecido , Lasers
11.
Sci Rep ; 14(1): 5753, 2024 03 08.
Artigo em Inglês | MEDLINE | ID: mdl-38459096

RESUMO

Parasitic organisms pose a major global health threat, mainly in regions that lack advanced medical facilities. Early and accurate detection of parasitic organisms is vital to saving lives. Deep learning models have uplifted the medical sector by providing promising results in diagnosing, detecting, and classifying diseases. This paper explores the role of deep learning techniques in detecting and classifying various parasitic organisms. The research works on a dataset consisting of 34,298 samples of parasites such as Toxoplasma Gondii, Trypanosome, Plasmodium, Leishmania, Babesia, and Trichomonad along with host cells like red blood cells and white blood cells. These images are initially converted from RGB to grayscale followed by the computation of morphological features such as perimeter, height, area, and width. Later, Otsu thresholding and watershed techniques are applied to differentiate foreground from background and create markers on the images for the identification of regions of interest. Deep transfer learning models such as VGG19, InceptionV3, ResNet50V2, ResNet152V2, EfficientNetB3, EfficientNetB0, MobileNetV2, Xception, DenseNet169, and a hybrid model, InceptionResNetV2, are employed. The parameters of these models are fine-tuned using three optimizers: SGD, RMSprop, and Adam. Experimental results reveal that when RMSprop is applied, VGG19, InceptionV3, and EfficientNetB0 achieve the highest accuracy of 99.1% with a loss of 0.09. Similarly, using the SGD optimizer, InceptionV3 performs exceptionally well, achieving the highest accuracy of 99.91% with a loss of 0.98. Finally, applying the Adam optimizer, InceptionResNetV2 excels, achieving the highest accuracy of 99.96% with a loss of 0.13, outperforming other optimizers. The findings of this research signify that using deep learning models coupled with image processing methods generates a highly accurate and efficient way to detect and classify parasitic organisms.


Assuntos
Babesia , Aprendizado Profundo , Parasitos , Toxoplasma , Animais , Microscopia
12.
Theranostics ; 14(5): 1794-1814, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38505609

RESUMO

Rationale: The acoustic stimulation of microbubbles within microvessels can elicit a spectrum of therapeutically relevant bioeffects from permeabilization to perfusion shutdown. These bioeffects ultimately arise from complex interactions between microbubbles and microvascular walls, though such interactions are poorly understood particularly at high pressure, due to a paucity of direct in vivo observations. The continued development of focused ultrasound methods hinges in large part on establishing links between microbubble-microvessel interactions, cavitation signals, and bioeffects. Methods: Here, a system was developed to enable simultaneous high-speed intravital imaging and cavitation monitoring of microbubbles in vivo in a chorioallantoic membrane model. Exposures were conducted using the clinical agent DefinityTM under conditions previously associated with microvascular damage (1 MHz, 0.5-3.5 MPa, 5 ms pulse length). Results: Ultrasound-activated microbubbles could be observed and were found to induce localized wall deformations that were more pronounced in smaller microvessels and increased with pressure. A central finding was that microbubbles could extravasate from microvessels (from 34% of vessels at 1 MPa to 79% at 3 MPa) during insonation (94% within 0.5 ms) and that this occurred more frequently and in progressively larger microvessels (up to 180 µm) as pressure was increased. Following microbubble extravasation, transient or sustained red blood cell leakage ensued at the extravasation site in 96% of cases for pressures ≥1 MPa. Conclusions: The results here represent the first high-speed in vivo investigation of high-pressure focused ultrasound-induced microbubble-microvessel interactions. This data provides direct evidence that the process of activated microbubble extravasation can occur in vivo and that it is linked to producing microvessel wall perforations of sufficient size to permit red blood cell leakage. The association of red blood cell leakage with microbubble extravasation provides mechanistic insight into the process of microvessel rupture, which has been widely observed in histology.


Assuntos
Membrana Corioalantoide , Microbolhas , Animais , Microscopia , Ultrassonografia/métodos , Microscopia Intravital
13.
Elife ; 122024 Mar 15.
Artigo em Inglês | MEDLINE | ID: mdl-38488831

RESUMO

Nondestructive pathology based on three-dimensional (3D) optical microscopy holds promise as a complement to traditional destructive hematoxylin and eosin (H&E) stained slide-based pathology by providing cellular information in high throughput manner. However, conventional techniques provided superficial information only due to shallow imaging depths. Herein, we developed open-top two-photon light sheet microscopy (OT-TP-LSM) for intraoperative 3D pathology. An extended depth of field two-photon excitation light sheet was generated by scanning a nondiffractive Bessel beam, and selective planar imaging was conducted with cameras at 400 frames/s max during the lateral translation of tissue specimens. Intrinsic second harmonic generation was collected for additional extracellular matrix (ECM) visualization. OT-TP-LSM was tested in various human cancer specimens including skin, pancreas, and prostate. High imaging depths were achieved owing to long excitation wavelengths and long wavelength fluorophores. 3D visualization of both cells and ECM enhanced the ability of cancer detection. Furthermore, an unsupervised deep learning network was employed for the style transfer of OT-TP-LSM images to virtual H&E images. The virtual H&E images exhibited comparable histological characteristics to real ones. OT-TP-LSM may have the potential for histopathological examination in surgical and biopsy applications by rapidly providing 3D information.


Assuntos
Microscopia , Neoplasias , Masculino , Humanos , Microscopia/métodos , Corantes Fluorescentes , Pele , Amarelo de Eosina-(YS) , Imageamento Tridimensional/métodos
14.
Soft Matter ; 20(12): 2804-2811, 2024 Mar 20.
Artigo em Inglês | MEDLINE | ID: mdl-38446076

RESUMO

The peels of lime, lemon, pomelo and citron are investigated at macroscopic and microscopic level. The structural composition of the peels is compared and properties such as peel thickness, proportion of flavedo, density and proportion of intercellular spaces are determined. µCT images are used to visualize vascular bundles and oil glands. SEM images provide information about the appearance of the cellular tissue in the outer flavedo and inner albedo. The proportion of intercellular spaces is quantitatively determined by manual and software-assisted analysis (ilastik). While there are macroscopic differences in the fruits, they differ only slightly in the orientation of the vascular bundles and the arrangement of the oil glands. However, in peel thickness and flavedo thickness, the fruit peels differ significantly from each other. There are no significant differences between the two analysis methods used, although the use of ilastik is preferred due to time reduction of up to 70%. The large amount of intercellular spaces in the albedo but also the denser flavedo both have a mechanical protective function to prevent damage to the fruit. In addition, the entire peel structure is mechanically reinforced by vascular bundles. This combination of penetration protection (flavedo) and energy dissipation (albedo) makes Citrus spp. peels a promising inspiration for technical material systems.


Assuntos
Citrus , Citrus/química , Citrus/ultraestrutura , Microscopia , Frutas/química , Frutas/ultraestrutura
15.
PLoS One ; 19(3): e0299875, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38498588

RESUMO

The widespread availability and diversity of open-source microcontrollers paired with off-the-shelf electronics and 3D printed technology has led to the creation of a wide range of low-cost scientific instruments, including microscopes, spectrometers, sensors, data loggers, and other tools that can be used for research, education, and experimentation. These devices can be used to explore a wide range of scientific topics, from biology and chemistry to physics and engineering. In this study, we designed and built a multifunction fluorescent open source in vivo/in vitro imaging system (openIVIS) system that integrates a Raspberry Pi with commercial cameras and LEDs with 3D printed structures combined with an acrylic housing. Our openIVIS provides three excitation wavelengths of 460 nm, 520 nm, and 630 nm integrated with Python control software to enable fluorescent measurements across the full visible light spectrum. To demonstrate the potential applications of our system, we tested its performance against a diverse set of experiments including laboratory assays (measuring fluorescent dyes, using optical nanosensors, and DNA gel electrophoresis) to potentially fieldable applications (plant and mineral imaging). We also tested the potential use for a high school biology environment by imaging small animals and tracking their development over the course of ten days. Our system demonstrated its ability to measure a wide dynamic range fluorescent response from millimolar to picomolar concentrations in the same sample while measuring responses across visible wavelengths. These results demonstrate the power and flexibility of open-source hardware and software and how it can be integrated with customizable manufacturing to create low-cost scientific instruments with a wide range of applications. Our study provides a promising model for the development of low-cost instruments that can be used in both research and education.


Assuntos
Eletrônica , Microscopia , Animais , Luz , Software , Tecnologia
16.
Curr Opin Cell Biol ; 87: 102342, 2024 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-38428224

RESUMO

Lipid droplets (LDs), once considered mere storage depots for lipids, have gained recognition for their intricate roles in cellular processes, including metabolism, membrane trafficking, and disease states like obesity and cancer. This review explores label-free imaging techniques' applications in LD research. We discuss holotomography and vibrational spectroscopic microscopy, emphasizing their potential for studying LDs without molecular labels, and we highlight the growing integration of artificial intelligence. Clinical applications in disease diagnosis and therapy are also considered.


Assuntos
Inteligência Artificial , Gotículas Lipídicas , Gotículas Lipídicas/metabolismo , Microscopia , Metabolismo dos Lipídeos
17.
Sci Rep ; 14(1): 5812, 2024 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-38461279

RESUMO

The increasing global demand for food, coupled with concerns about the environmental impact of synthetic fertilizers, underscores the urgency of developing sustainable agricultural practices. Nitrogen-fixing bacteria, known as diazotrophs, offer a potential solution by converting atmospheric nitrogen into bioavailable forms, reducing the reliance on synthetic fertilizers. However, a deeper understanding of their interactions with plants and other microbes is needed. In this study, we introduce a recently developed label-free 3D quantitative phase imaging technology called dynamic quantitative oblique back-illumination microscopy (DqOBM) to assess the functional dynamic activity of diazotrophs in vitro and in situ. Our experiments involved three different diazotrophs (Sinorhizobium meliloti, Azotobacter vinelandii, and Rahnella aquatilis) cultured on media with amendments of carbon and nitrogen sources. Over 5 days, we observed increased dynamics in nutrient-amended media. These results suggest that the observed bacterial dynamics correlate with their metabolic activity. Furthermore, we applied qOBM to visualize microbial dynamics within the root cap and elongation zone of Arabidopsis thaliana primary roots. This allowed us to identify distinct areas of microbial infiltration in plant roots without the need for fluorescent markers. Our findings demonstrate that DqOBM can effectively characterize microbial dynamics and provide insights into plant-microbe interactions in situ, offering a valuable tool for advancing our understanding of sustainable agriculture.


Assuntos
Arabidopsis , Fertilizantes , Fertilizantes/microbiologia , Iluminação , Microscopia , Plantas/metabolismo , Arabidopsis/metabolismo , Nitrogênio/metabolismo , Fixação de Nitrogênio
18.
Proc Natl Acad Sci U S A ; 121(12): e2304866121, 2024 Mar 19.
Artigo em Inglês | MEDLINE | ID: mdl-38483992

RESUMO

Accelerating the measurement for discrimination of samples, such as classification of cell phenotype, is crucial when faced with significant time and cost constraints. Spontaneous Raman microscopy offers label-free, rich chemical information but suffers from long acquisition time due to extremely small scattering cross-sections. One possible approach to accelerate the measurement is by measuring necessary parts with a suitable number of illumination points. However, how to design these points during measurement remains a challenge. To address this, we developed an imaging technique based on a reinforcement learning in machine learning (ML). This ML approach adaptively feeds back "optimal" illumination pattern during the measurement to detect the existence of specific characteristics of interest, allowing faster measurements while guaranteeing discrimination accuracy. Using a set of Raman images of human follicular thyroid and follicular thyroid carcinoma cells, we showed that our technique requires 3,333 to 31,683 times smaller number of illuminations for discriminating the phenotypes than raster scanning. To quantitatively evaluate the number of illuminations depending on the requisite discrimination accuracy, we prepared a set of polymer bead mixture samples to model anomalous and normal tissues. We then applied a home-built programmable-illumination microscope equipped with our algorithm, and confirmed that the system can discriminate the sample conditions with 104 to 4,350 times smaller number of illuminations compared to standard point illumination Raman microscopy. The proposed algorithm can be applied to other types of microscopy that can control measurement condition on the fly, offering an approach for the acceleration of accurate measurements in various applications including medical diagnosis.


Assuntos
Microscopia , Análise Espectral Raman , Humanos , Microscopia/métodos , Análise Espectral Raman/métodos , Glândula Tireoide , Microscopia Óptica não Linear , Aprendizado de Máquina
19.
Sci Rep ; 14(1): 6662, 2024 03 20.
Artigo em Inglês | MEDLINE | ID: mdl-38509285

RESUMO

Acute lung injury (ALI) represents an aetiologically diverse form of pulmonary damage. Part of the assessment and diagnosis of ALI depends on skilled observer-based scoring of brightfield microscopy tissue sections. Although this readout is sufficient to determine gross alterations in tissue structure, its categorical scores lack the sensitivity to describe more subtle changes in lung morphology. To generate a more sensitive readout of alveolar perturbation we carried out high resolution immunofluorescence imaging on 200 µm lung vibratome sections from baseline and acutely injured porcine lung tissue, stained with a tomato lectin, Lycopersicon Esculentum Dylight-488. With the ability to resolve individual alveoli along with their inner and outer wall we generated continuous readouts of alveolar wall thickness and circularity. From 212 alveoli traced from 10 baseline lung samples we established normal distributions for alveolar wall thickness (27.37; 95% CI [26.48:28.26]) and circularity (0.8609; 95% CI [0.8482:0.8667]) in healthy tissue. Compared to acutely injured lung tissue baseline tissue exhibited a significantly lower wall thickness (26.86 ± 0.4998 vs 50.55 ± 4.468; p = 0.0003) and higher degree of circularityϕ≤ (0.8783 ± 0.01965 vs 0.4133 ± 0.04366; p < 0.0001). These two components were subsequently combined into a single more sensitive variable, termed the morphological quotient (MQ), which exhibited a significant negative correlation (R2 = 0.9919, p < 0.0001) with the gold standard of observer-based scoring. Through the utilisation of advanced light imaging we show it is possible to generate sensitive continuous datasets describing fundamental morphological changes that arise in acute lung injury. These data represent valuable new analytical tools that can be used to precisely benchmark changes in alveolar morphology both in disease/injury as well as in response to treatment/therapy.


Assuntos
Lesão Pulmonar Aguda , Pulmão , Animais , Suínos , Alvéolos Pulmonares/diagnóstico por imagem , Lesão Pulmonar Aguda/diagnóstico por imagem , Microscopia , Imagem Óptica
20.
Nanoscale ; 16(12): 6190-6198, 2024 Mar 21.
Artigo em Inglês | MEDLINE | ID: mdl-38445876

RESUMO

Here we introduce scattering-type scanning near-field optical microscopy (s-SNOM) as a novel tool for nanoscale chemical-imaging of sub-cellular organelles, nanomaterials and of the interactions between them. Our setup uses a tuneable mid-infrared laser and a sharp scanning probe to image at a resolution substantially surpassing the diffraction limit. The laser can be tuned to excite vibrational modes of functional groups in biomolecules, (e.g. amide moieties), in a way that enables direct chemical mapping without the need for labelling. We, for the first time, chemically image neuronal ultrastructure, identify neuronal organelles and sub-organelle structures as small as 10 nm and validate our findings using transmission electron microscopy (TEM). We produce chemical and morphological maps of neurons treated with gold nanospheres and characterize nanoparticle size and intracellular location, and their interaction with the plasma membrane. Our results show that the label-free nature of s-SNOM means it has a 'true' chemical resolution of up to 20 nm which can be further improved. We argue that it offers significant potential in nanomedicine for nanoscale chemical imaging of cell ultrastructure and the subcellular distribution of nanomaterials within tissues.


Assuntos
Nanopartículas , Nanoestruturas , Nanotecnologia/métodos , Microscopia/métodos , Nanoestruturas/química , Luz
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