We present a multimodal technique for measuring the integral refractive index and the thickness of biological cells and their organelles by integrating interferometric phase microscopy (IPM) and rapid confocal fluorescence microscopy. First, the actual thickness maps of the cellular compartments are reconstructed using the confocal fluorescent sections, and then the optical path difference (OPD) map of the same cell is reconstructed using IPM. Based on the co‐registered data, the integral refractive index maps of the cell and its organelles are calculated. This technique enables rapidly measuring refractive index of live, dynamic cells, where IPM provides quantitative imaging capabilities and confocal fluorescence microscopy provides molecular specificity of the cell organelles. We acquire human colorectal adenocarcinoma cells and show that the integral refractive index values are similar for the whole cell, the cytoplasm and the nucleus on the population level, but significantly different on the single cell level. 相似文献
Platinum chemosensitivity detection plays a vital role during endometrial cancer treatment because chemotherapy responses have profound influences on patient's prognosis. Although several methods can be used to detect drug resistance characteristics, studies on detecting drug sensitivity based on dynamic and quantitative phase imaging of cancer cells are rare. In this study, digital holographic microscopy was applied to distinguish drug‐resistant and nondrug‐resistant endometrial cancer cells. Based on the reconstructed phase images, temporal evolutions of cell height (CH), cell projected area (CPA) and cell volume were quantitatively measured. The results show that change rates of CH and CPA were significantly different between drug‐resistant and nondrug‐resistant endometrial cancer cells. Furthermore, the results demonstrate that morphological characteristics have the potential to be utilized to distinguish the drug sensitivity of endometrial cancer cells, and it may provide new perspectives to establish optical methods to detect drug sensitivity and guide chemotherapy in endometrial cancer. 相似文献
Melasma is a skin disorder characterized by hyperpigmented patches due to increased melanin production and deposition. In this pilot study, we evaluate the potential of multiphoton microscopy (MPM) to characterize non‐invasively the melanin content, location, and distribution in melasma and assess the elastosis severity. We employed a clinical MPM tomograph to image in vivo morphological features in melasma lesions and adjacent normal skin in 12 patients. We imaged dermal melanophages in most dermal melasma lesions and occasionally in epidermal melasma. The melanin volume fraction values measured in epidermal melasma (14% ± 4%) were significantly higher (p < 0.05) than the values measured in perilesional skin (11% ± 3%). The basal keratinocytes of melasma and perilesions showed different melanin distribution. Elastosis was predominantly more severe in lesions than in perilesions and was associated with changes in melanin distribution of the basal keratinocytes. These results demonstrate that MPM may be a non‐invasive imaging tool for characterizing melasma. 相似文献
The advent of super‐resolution microscopy allowed for new insights into cellular and physiological processes of normal and diseased cells. In this study, we report for the first time on the super‐resolved DNA structure of buccal cells from patients with Alzheimer's disease (AD) versus age‐ and gender‐matched healthy, non‐caregiver controls. In this super‐resolution study cohort of 74 participants, buccal cells were collected and their spatial DNA organization in the nucleus examined by 3D Structured Illumination Microscopy (3D‐SIM). Quantitation of the super‐resolution DNA structure revealed that the nuclear super‐resolution DNA structure of individuals with AD significantly differs from that of their controls (p < 0.05) with an overall increase in the measured DNA‐free/poor spaces. This represents a significant increase in the interchromatin compartment. We also find that the DNA structure of AD significantly differs in mild, moderate, and severe disease with respect to the DNA‐containing and DNA‐free/poor spaces. We conclude that whole genome remodeling is a feature of buccal cells in AD. 相似文献
1 The present study used a crop life table to determine the critical components of production and the key factors of loss in tomato, and three treatments to identify the integrated pest management (IPM) benefits on the reduction of yield losses and the conservation of natural enemies. 2 The relative IPM benefits were compared using calendar‐based pesticide applications, IPM and control (no pesticide). A total of 1248 tomato plants were allotted to treatments with four replicates of 104 plants, each in a random block design. The densities of vectors, leaf miners, fruit borers, predators and parasitoids were compared. 3 Fruit was the critical component of production, experiencing the greatest losses, followed by flower and plant in the vegetative phase. The key causes of loss of production were tospoviruses, Erwinia carotovora, Alternaria solani, Phytophthora infestans, Neoleucinodes elegantalis and blossom‐end rot. 4 No significant differences in yield were detected between the calendar‐based and IPM systems. In the control, the yield was lower than the yield in treatments with pesticides due to losses from fungal diseases and viruses. IPM more efficiently controlled pests than the calendar‐system, reducing the number of parathion‐methyl and abamectin applications by 3.8‐ and 2.9‐fold, respectively. IPM treatment significantly reduced the impact of pesticides on natural enemies. 5 Tomato yield was more affected by biotic and abiotic factors during the reproductive stage. Because fruit was the production component most susceptible to loss, cultivation and IPM programmess should prioritize practices to reduce loss of this component. 相似文献
We present a dual‐modality technique based on wide‐field photothermal (PT) interferometric phase imaging and simultaneous PT ablation to selectively deplete specific cell populations labelled by plasmonic nanoparticles. This combined technique utilizes the plasmonic reaction of gold nanoparticles under optical excitation to produce PT imaging contrast by inducing local phase changes when the excitation power is weak, or ablation of selected cells when increasing the excitation power. Controlling the entire process is carried out by dynamic quantitative phase imaging of all cells (labelled and unlabelled). We demonstrate our ability to detect and specifically ablate in vitro cancer cells over‐expressing epidermal growth factor receptors (EGFRs), labelled with plasmonic nanoparticles, in the presence of either EGFR under‐expressing cancer cells or white blood cells. The latter demonstration establishes an initial model for depletion of circulating tumour cells in blood. The proposed system is able to image in wide field the label‐free quantitative phase profile together with the PT phase profile of the sample, and provides the ability of both detection and selective cell ablation in a controlled environment.
Quantitative phase imaging with molecular specificity and specific cell depletion. ( a ) Label‐free quantitative phase profiles of mixed population of EGFR+/EGFR– cancer cells. ( b ) When weak modulated PT excitation is applied, selective phase contrast is generated in the modulation frequency only for the EGFR+ cancer cells labelled with plasmonic nanoparticles. ( c ) When stronger modulated PT excitation is applied, selective ablation of the EGFR+ cancer cells labelled with plasmonic nanoparticles occurs. White scalebars represent 10 µm upon sample. 相似文献
Recently, beauvericin (BEA) has been recognized as an important toxic compound synthesized by several Fusarium strains, infecting maize, wheat, and rice, worldwide. The effects of BEA on mammalian cells have been studied; however, its effects on the function of host plant cells are largely unknown. The purpose of our work was to assess whether BEA can affect the root and leaf cells of wheat cultivar (cv.) ‘Arina’ seedlings, using a cytotoxicity assay and fluorescence microscopy. Toxigenicity during wheat germination was higher in BEA‐treated wheat seedlings than in non‐treated seedlings (control). Leaf primordial, situated at the base and the tips of treated leaves, were more affected by BEA compared to the control when assayed in medium for cell viability measured by luminescent equipment. BEA‐Treated plant cells secrete adenosine triphosphate (ATP) to the extracellular matrix and invoke more luminescence by luciferase than the non‐treated seedlings. Our results were confirmed by fluorescence microscopy following ‘4′,6‐diamidino‐2‐phenylindole’ (DAPI) staining and by confocal microscopy. In addition, the bioluminescent protein luciferase was observed in the intracellular space indicating presence of ATP. The incidence of nuclear fragmentation increased significantly in cells of seedlings treated with BEA at 40 μM concentration implying that the intracellular phytotoxin BEA plays an important role, possibly as a mediator in cell‐death signalling. 相似文献
Cancer initiating cells (CICs) have been the focus of recent anti‐cancer therapies, exhibiting strong invasion capability via potentially enhanced ability to remodel extracellular matrices (ECM). We have identified CICs in a human breast cancer cell line, MX‐1, and developed a xenograft model in SCID mice. We investigated the CICs' matrix‐remodeling effects using Second Harmonic Generation (SHG) microscopy to identify potential phenotypic signatures of the CIC‐rich tumors. The isolated CICs exhibit higher proliferation, drug efflux and drug resistant properties in vitro; were more tumorigenic than non‐CICs, resulting in more and larger tumors in the xenograft model. The CIC‐rich tumors have less collagen in the tumor interior than in the CIC‐poor tumors supporting the idea that the CICs can remodel the collagen more effectively. The collagen fibers were preferentially aligned perpendicular to the CIC‐rich tumor boundary while parallel to the CIC‐poor tumor boundary suggesting more invasive behavior of the CIC‐rich tumors. These findings would provide potential translational values in quantifying and monitoring CIC‐rich tumors in future anti‐cancer therapies.
CIC‐rich tumors remodel the collagen matrix more than CIC‐poor tumors. 相似文献
Electron microscopy (EM) has been a key imaging method to investigate biological ultrastructure for over six decades. In recent years, novel volume EM techniques have significantly advanced nanometre‐scale imaging of cells and tissues in three dimensions. Previously, this had depended on the slow and error‐prone manual tasks of cutting and handling large numbers of sections, and imaging them one‐by‐one with transmission EM. Now, automated volume imaging methods mostly based on scanning EM (SEM) allow faster and more reliable acquisition of serial images through tissue volumes and achieve higher z‐resolution. Various software tools have been developed to manipulate the acquired image stacks and facilitate quantitative analysis. Here, we introduce three volume SEM methods: serial block‐face electron microscopy (SBEM), focused ion beam SEM (FIB‐SEM) and automated tape‐collecting ultramicrotome SEM (ATUM‐SEM). We discuss and compare their capabilities, provide an overview of the full volume SEM workflow for obtaining 3D datasets and showcase different applications for biological research. 相似文献
The purpose of this paper was to study the effect of the isopropyl myristic acid ester (IPM) on the physicochemical characteristics
of etoposide-loaded poly(lactic-co-glycolic acid) (PLGA) microspheres-specifically, the effects on the size and drug loading
of the microspheres, the polymer matrix and surface morphology, and the release of etoposide from the microspheres. The experiment
was structured to examine 2 IPM concentrations (25% and 50%) and 1 control (no IPM) at 2 different etoposide-loading percentages
(10% and 5%). The microspheres were prepared using a single-emulsion solvent-extraction procedure. Samples from each batch
of microspheres were then analyzed for size distribution. drug-loading efficiency, surface characteristics, in vitro release,
and in vitro microsphere degradation. The incorporation of 50% IPM significantly increased (P<05) the size of the microspheres when compared with the control and 25% IPM microspheres. However, incorporation of 25% or
50% IPM did not change (P>.05) the drug-loading efficiency in comparison with the microspheres prepared without IPM. The microspheres containing 50%
IPM were shown to significantly increase (P<.05) the release of etoposide from the microspheres at both etoposide concentrations. The microspheres prepared incorporating
25% IPM and 5% etoposide increased the in vitro release (P<.05) in comparison with the microspheres prepared without IPM. The 5% etoposide-PLGA microspheres showed a smooth, nonporous
surface that changed to a dimpled. nonporous surface after addition of 25% IPM. During the in vitro degradation study, the
IPM-containing microspheres slowly became porous but retained their structural integrity throughout the experiment. 相似文献
Recently developed super‐resolution microscopy techniques are changing our understanding of lipid rafts and membrane organisation in general. The lipid raft hypothesis postulates that cholesterol can drive the formation of ordered domains within the plasma membrane of cells, which may serve as platforms for cell signalling and membrane trafficking. There is now a wealth of evidence for these domains. However, their study has hitherto been hampered by the resolution limit of optical microscopy, making the definition of their properties problematic and contentious. New microscopy techniques circumvent the resolution limit and, for the first time, allow the fluorescence imaging of structures on length scales below 200 nm. This review describes such techniques, particularly as applied to the study of membrane organisation, synthesising newly emerging facets of lipid raft biology into a state‐of‐the art model. Editor's suggested further reading in BioEssays: Super‐resolution imaging prompts re‐thinking of cell biology mechanisms Abstract and Quantitative analysis of photoactivated localization microscopy (PALM) datasets using pair‐correlation analysis Abstract 相似文献
Photoacoustic microscopy (PAM) provides a fundamentally new tool for a broad range of studies of biological structures and functions. However, the use of PAM has been largely limited to small vertebrates due to the large size/weight and the inconvenience of the equipment. Here, we describe a portable optical‐resolution photoacoustic microscopy (pORPAM) system for 3‐dimensional (3D) imaging of small‐to‐large rodents and humans with a high spatiotemporal resolution and a large field of view. We show extensive applications of pORPAM to multiscale animals including mice and rabbits. In addition, we image the 3D vascular networks of human lips, and demonstrate the feasibility of pORPAM to observe the recovery process of oral ulcer and cancer‐associated capillary loops in human oral cavities. This technology is promising for broad biomedical studies from fundamental biology to clinical diseases. 相似文献
Adipogenesis and increase in fat tissue mass are mechanosensitive processes and hence should be influenced by the mechanical properties of adipocytes. We evaluated subcellular effective stiffnesses of adipocytes using atomic force microscopy (AFM) and interferometric phase microscopy (IPM), and we verified the empirical results using finite element (FE) simulations. In the AFM studies, we found that the mean ratio of stiffnesses of the lipid droplets (LDs) over the nucleus was 0.83 ± 0.14, from which we further evaluated the ratios of LDs over cytoplasm stiffness, as being in the range of 2.5 to 8.3. These stiffness ratios, indicating that LDs are stiffer than cytoplasm, were verified by means of FE modeling, which simulated the AFM experiments, and provided good agreement between empirical and model-predicted structural behavior. In the IPM studies, we found that LDs mechanically distort their intracellular environment, which again indicated that LDs are mechanically stiffer than the surrounding cytoplasm. Combining these empirical and simulation data together, we provide in this study evidence that adipocytes stiffen with differentiation as a result of accumulation of LDs. Our results are relevant to research of adipose-related diseases, particularly overweight and obesity, from a mechanobiology and cellular mechanics perspectives. 相似文献