Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “Image Cytometry”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,369 records · Page 76Linked to original sources
PubMed2006

Cytoarchitectonic identification and probabilistic mapping of two distinct areas within the anterior ventral bank of the human intraparietal sulcus.

Anatomical studies in the macaque cortex and functional imaging studies in humans have demonstrated the existence of different cortical areas within the intraparietal sulcus (IPS). Such functional segregation, however, does not correlate with presently available architectonic maps of the human brain. This is particularly true for the classical Brodmann map, which is still widely used as an anatomical reference in functional imaging studies. The aim of this cytoarchitectonic mapping study was to use previously defined algorithms to determine whether consistent regions and borders can be found within the cortex of the anterior IPS in a population of 10 post-mortem human brains. Two areas, the human intraparietal area 1 (hIP1) and the human intraparietal area 2 (hIP2), were delineated in serial histological sections of the anterior, lateral bank of the human IPS. The region hIP1 is located posterior and medial to hIP2, and the former is always within the depths of the IPS. The latter, on the other hand, sometimes reaches the free surface of the superior parietal lobule. The delineations were registered to standard reference space, and probabilistic maps were calculated, thereby quantifying the intersubject variability in location and extent of both areas. In the future, they can be a tool for analyzing structure-function relationships and a basis for determining degrees of homology in the IPS among anthropoid primates. We conclude that the human IPS has a more finely grained parcellation than shown in Brodmann's map.

Adult↗
PubMed2006

Ontogeny of 5-HT neurons in the brainstem of the lamprey, Petromyzon marinus.

This study examined the spatial and temporal distribution of serotonin-immunoreactive (5-HT-ir) neurons in the brainstem of Petromyzon marinus at three developmental stages, larval, postmetamorphic, and reproductive. Computer-assisted 3-D reconstructions were made of the three main 5-HT-ir neuron groups. The rostralmost brainstem group was located near the posterior commissure, the second group at the isthmus, and the third group in the bulbar area. For each of those groups, the distribution of the 5-HT-ir neurons was very similar in the three developmental stages examined, suggesting that the 5-HT system is relatively mature early in larval animals. The soma of 5-HT-ir neurons increased in size and their dendritic fields increased in complexity with development. Furthermore, the number of 5-HT-ir neurons in each group increased significantly from the larval to the reproductive stage. To determine whether this was due to the genesis of 5-HT neurons, bromodeoxyuridine (BrdU) was injected into larval, metamorphosing, and postmetamorphic lampreys. These experiments revealed a few neurons colocalizing BrdU and 5-HT in metamorphosing animals. Taken together, the present results suggest that 5-HT neurons increase in number during maturation and that neurogenesis could, at least partially, contribute to the appearance of new 5-HT cells at different developmental stages.

Animals↗
PubMed2006

Model-based analysis of excitatory lateral connections in the visual cortex.

Excitatory lateral connections within the primary visual cortex are thought to link neurons with similar receptive field properties. Here we studied whether this rule can predict the distribution of excitatory connections in relation to cortical location and orientation preference in the cat visual cortex. To this end, we obtained orientation maps of areas 17 or 18 using optical imaging and injected anatomical tracers into these regions. The distribution of labeled axonal boutons originating from large populations of excitatory neurons was then analyzed and compared with that of individual pyramidal or spiny stellate cells. We demonstrate that the connection patterns of populations of nearby neurons can be reasonably predicted by Gaussian and von Mises distributions as a function of cortical location and orientation, respectively. The connections were best described by superposition of two components: a spatially extended, orientation-specific and a local, orientation-invariant component. We then fitted the same model to the connections of single cells. The composite pattern of nine excitatory neurons (obtained from seven different animals) was consistent with the assumptions of the model. However, model fits to single cell axonal connections were often poorer and their estimated spatial and orientation tuning functions were highly variable. We conclude that the intrinsic excitatory network is biased to similar cortical locations and orientations but it is composed of neurons showing significant deviations from the population connectivity rule.

Animals↗
PubMed2003

Chronology of cellular alterations during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells: analysis by time lapse-video microscopy and conventional fluorescence microscopy.

BACKGROUND: Time-lapse video microscopy was used to determine whether mitochondrial and nuclear changes (decrease in mitochondrial transmembrane potential, condensation, and/or fragmentation of the nuclei, morphologic features typical of apoptosis) occurring during 7-ketocholesterol-induced cell death on A7R5 rat smooth muscle cells took place before or after the loss of cell adhesion. In addition, changes in actin organization were followed by conventional fluorescence microscopy. METHODS: Morphologic, functional, and spatial changes at the mitochondrial level were investigated with 3,3'-dihexyloxacarbocyanine iodide and/or MitoTracker Red, and nuclear morphology was characterized by staining with Hoechst 33342. Actin fibers, which are major components of the filament network of the cytoskeleton, were visualized with phalloidin linked to fluorescein. The numbers of adherent and nonadherent cells were determined by cell counting. RESULTS: 7-Ketocholesterol-induced cell death was associated with a rapid alteration of actin fibers, a loss of intercellular junctions, and cell shape modifications. Analysis of mitochondrial transmembrane potential showed successively a hyperpolarization and a more or less pronounced progressive decrease followed by a dramatic drop associated with an increase in Hoechst 33342 staining, reflecting chromatin condensation and morphologic changes in the nuclei. CONCLUSIONS: During cell death induced by 7-ketocholesterol in A7R5 rat smooth muscle cells, the different methods of microscopy allowed us to establish that alterations of actin fibers and mitochondrial dysfunctions occurred before condensation and/or fragmentation of the nuclei, which preceded the loss of cell adhesion.

Actins↗
PubMed2003

Fluorescence lifetime-resolved pH imaging of living cells.

BACKGROUND: The regulation and maintenance of intracellular pH are critical to diverse metabolic functions of the living cells. Fluorescence time-resolved techniques and instrumentations have advanced rapidly and enabled the imaging of intracellular pH based on the fluorescence lifetimes. METHODS: The frequency-domain fluorescence lifetime imaging microscopy (FLIM) and fluorophores displaying appropriate pH-dependent lifetime sensitivities were used to determine the temporal and spatial pH distributions in the cytosol and vesicular compartment lysosomes. RESULTS: We found that cytosolic pH levels are different in 3T3 fibroblasts, Chinese hamster ovary (CHO) cells, and MCF-7 cells when using the pH probe carboxy-SNAFL2. We also tracked the transient cytosolic pH changes in the living CHO cells after treatments with proton pump inhibitors, ion exchanger inhibitors, and weak base and acid. The intracellular lysosomal pH was determined with the acidic lifetime probes DM-NERF dextrans, OG-514 carboxylic acid dextrans, and LysoSensor DND-160. Our results showed that the resting lysosomal pH value obtained from the 3T3 fibroblasts was between 4.5 and 4.9. The increase of lysosomal pH induced by the treatments with proton pump inhibitor and ionophores also were observed in our FLIM measurements. CONCLUSIONS: Our lifetime-based pH imaging data suggested that FLIM can measure the intracellular pH of the resting cells and follow the pH fluctuations inside the cells after environmental perturbations. To improve the z-axis resolution to the intracellular lifetime-resolved images, we are investigating the implementation of the pseudo-confocal capability to our current FLIM apparatus.

3T3 Cells↗
PubMed2003

DNA content, chromatin texture and nuclear morphology in benzo[a]pyrene-transformed human breast epithelial cells after microcell-mediated transfer of chromosomes 11 and 17.

BACKGROUND: A relation between the changes in DNA content and chromatin supra-organization and the expression of gradual steps of tumorigenesis has been assessed by image analysis in human breast epithelial cells (MCF-10F) treated with benzo[a]pyrene (BP) (cell lines BP1, BP1-E, BP1-Tras, and others). METHODS: Because abnormal chromosomes 11 and 17 have been associated with neoplastic progression in BP-transformed MCF-10F cells, image analysis of Feulgen-stained tumorigenic BP1-E cells with the microcell-mediated chromosome transfer of normal chromosomes 11 and 17 was carried out. RESULTS: A tendency of DNA amount distribution and nuclear size restoration to values typical of non-transformed MCF-10F cells was demonstrated, especially after the transfer of chromosome 17. No reversion in chromatin texture was found after the transfer of chromosome 11 or 17. CONCLUSIONS: Although the presence of a normal chromosome 17 should be considered among the necessary steps for tumorigenic human breast epithelial cells to recover their normality, a more complex genome balance is required for the entire nuclear chromatin of these cells to recover its totally normal supra-organization and expression.

Benzo(a)pyrene↗
PubMed2003

Characterization of dynamic cellular adhesion of osteoblasts using atomic force microscopy.

BACKGROUND: Atomic force microscopy (AFM) can be used to visualize the cell morphology in an aqueous environment and in real time. It also allows the investigation of mechanical properties such as cell compliance as a function of cell attachment. This study characterized and evaluated osteoblast adhesion by AFM. METHODS: Human bone marrow stromal cells were cultured on two types of surface to induce weak and strong cellular adhesions. RESULTS: Cells were considered as spreading if they had a flattened and lengthened shape and a cytoskeletal organization in the submembrane cytosolic region. Cell detachment demonstrated different adhesion states between adherent cells to be distinguished. The stability of the cytoskeletal fibers indicated that cells were adherent. The elastic modulus was estimated by two complementary approaches. The values deduced were between 3 x 10(2) and 2 x 10(5) Nm(-2) according to the state of cell adhesion and the approaches used to measure this elastic modulus. CONCLUSIONS: Although the results were qualitative, a relation may be deduced between the elasticity of living cells as demonstrated by cytoskeletal organization and the state of cell adhesion. The technique could be used to determine the adhesion state of an adherent osteoblast observed under AFM.

Bone Marrow Cells↗
PubMed2003

Quantitative analysis of cytoskeletal organization by digital fluorescent microscopy.

BACKGROUND: The cytoskeleton consists of complex arrays of fibers that play indispensable roles in cell structure and function. The cytoskeletal fibers are concertedly involved in numerous cellular processes, including cell adhesion, locomotion, intracellular transport, and cell division. The organization of the cytoskeleton was extensively studied, mainly by immunofluorescence microscopy, yet these studies were mostly qualitative, and a reliable quantitative approach for determining fiber structure and distribution is still missing. METHODS: In this study we developed algorithms for filament feature extraction, based on fluorescence microscopy. These algorithms are robust against blurring by slight defocus, high background, and noise, and are applicable to both fixed, immunolabeled cells and live cells expressing fluorescently tagged cytoskeletal proteins. The implemented FiberScore program is used in order to recognize, segment, and quantify various structural parameters of the cytoskeleton, including total fiber-associated fluorescence, as well as fiber length and orientation. Furthermore, these parameters can be determined for different cytoskeletal proteins in the same sample tagged with multiple-fluorescent labels, and the results can be correlated with other cellular parameters. RESULTS: FiberScore was used here for the quantification of simultaneous changes in microtubule and actin filaments induced by the microtubule-disrupting drug nocodazole. Actin filaments, which are reported to respond reciprocally to microtubule disruption, are found to be affected by both immediate and delayed signals. CONCLUSIONS: Analysis of the organization of fibers by the FiberScore algorithm allows quantification of the cytoskeletal signature of cells and offers reliable multiparametric functional assays for effects of drugs and other perturbations evaluated on a cell-by-cell basis.

3T3 Cells↗
PubMed2003

An analysis of changes in the expression of cyclins A and B1 by the cell array system during the cell cycle: comparison between cell synchronization methods.

BACKGROUND: A novel high-throughput analysis, cell array system, was developed for an extensive study of the expression of genes and/or the degradation of gene products at the cellular level. To exemplify the usefulness of this system, we showed the changes in the expression level of cyclin A and B1 during the cell cycle in a single experiment. METHODS: We used the cell array system to chase the changes in cyclin A and B1 expression during the cell cycle in HeLa cells. Cells were synchronized by mitotic selection and thymidine-hydroxyurea methods. Cells were harvested at intervals of 1 h from 0 through 23 h. These 48 cell samples were spotted on a circle of the cell array glass slide. Cyclin A and B1 were immunologically stained with Alexa Fluor 488, and nuclear DNA was stained with propidium iodide. The amount of cyclins and nuclear DNA were simultaneously measured by a laser scanning cytometer. RESULTS: Both cyclins were expressed in a cell cycle-dependent manner as previously reported. The precise time-course of the expression level of cyclins were obtained at a single experiment with this cell array system. CONCLUSIONS: This study indicates that the cell array system is valuable to analyze temporal course of protein expression in relation to the cell cycle position and, that it facilitates antigen expression studies at the cellular level in multiple samples.

Cell Cycle↗
PubMed2003

Lectinocytochemical detection of apoptotic murine leukemia L1210 cells.

BACKGROUND: Various cytomorphologic and biochemical markers of apoptosis are found in different compartments (plasma membrane, cytoplasm, nucleus, and mitochondria) of target cells. Although the plasma membrane is an easily accessible cellular compartment, relatively little is known about its changes during apoptosis. We investigated whether specific changes in the expression of plasma membrane glycoproteins take place during apoptosis and whether these changes could be used for a quantitative estimation of apoptosis. METHODS: Lectin cytochemical study of normal and apoptotic murine leukemia cells of the L1210 line was done. Horseradish peroxidase-labeled Laburnum anagyroides bark agglutinin, Phaseolus vulgaris agglutinin, Pisum sativum lectin (PSL), Ricinus communis agglutinin (RCA-120; 120 kDa), Solanum tuberosum agglutinin, Triticum vulgaris lectin (wheat germ agglutinin), Viscum album agglutinin, Canavalia ensiformis lectin (concanavalin A), and Helix pomatia lectin were used to compare specific glycoprotein expressions in normal and apoptotic murine leukemia cells of the L1210 line sensitive (L1210) and resistant (L1210R) to apoptosis induction by cisplatin. RESULTS: The data demonstrated significantly increased binding of alpha-D-mannose-specific PSL lectin (P<0.01) and beta-D-galactose-specific RCA-120 lectin (P<0.001) by the apoptotic cells of the L1210 and L1210R lines in comparison with the intact cells. That binding was shown to be specific because it was blocked by the corresponding inhibitory sugars. CONCLUSIONS: Lectins specific to alpha-D-mannose (PSL) and beta-D-galactose (RCA-120) can be used to distinguish between native and apoptotic murine leukemia L1210 cells and to quantitatively estimate apoptosis in a population of these cells.

Animals↗
PubMed2004

Fourier transformed spectral bio-imaging for studying the intracellular fate of liposomes.

BACKGROUND: To improve the efficiency of liposomal drug targeting systems, it is necessary to understand the mechanism of liposome uptake by the cell and to follow the intracellular fate of internalized liposomes and their contents. METHODS: We applied multiple-color fluorescence imaging spectroscopy, using a combination of five fluorescent dyes with a significant spectral overlap. pH-sensitive liposomes were labeled with the hydrophilic dye fluorescein isothiocyanate-dextran (FITC-dextran) or the lipophilic membrane marker rhodamine-B-phosphoethanolamine (Rh-PE) and incubated with COS-7 cells. Further, the cells were stained with specific markers: the cell membrane was fluorescently labeled with Vybrant DiO, lysosomes were stained with LysoTracker Red, and 4',6 diamidino-2-phenylindole dihydrochloride was used for counterstaining the nucleus. RESULTS: All five dyes were used simultaneously and were spectrally distinguished by the system. FITC-dextran-labeled liposomes showed a distribution pattern different from identically composed liposomes labeled with Rh-PE: the highly lipophilic Rh-PE was colocalized with the lysosomotropic dye LysoTracker Red, whereas liposomal FITC-dextran was not accompanied by LysoTracker Red in all cases. CONCLUSIONS: (a) Spectral (bio-) imaging is a powerful method for studying the intracellular fate of liposomal compounds. (b) We assume that the liposome membrane marker Rh-PE influences the uptake of particles due to its surface-modifying properties. We propose that this head-group-labeled phospholipid acts as a ligand for cellular receptors and triggers receptor-mediated (clathrin-dependent) endocytosis.

Algorithms↗
PubMed2004

Direct neural network application for automated cell recognition.

BACKGROUND: Automated cell recognition from histologic images is a very complex task. Traditionally, the image is segmented by some methods chosen to suit the image type, the objects are measured, and then a classifier is used to determine cell type from the object's measurements. Different classifiers have been used with reasonable success, including neural networks working with data from morphometric analysis. METHODS: Image data of cells were input directly into neural networks to determine the feasibility of direct classification by using pixel intensity information. Several types of neural network and their ability to work with cells in a complex patterned background were assessed for a variety of images and cell types and for the accuracy of classification. RESULTS: Inflammatory cells from animal biomaterial implants in rabbit paravertebral muscle were imaged in histologic sections. Simple, three-layer, fully connected, back-propagation neural networks and four-layer networks with two layers of a shared-weights neural network were most successful at classifying the cells from the images, with 97% and 98% correct recognition rates, respectively. CONCLUSIONS: The high accuracy recognition rate shows the potential for direct classification of visual image pixel data by neural networks.

Animals↗
PubMed2006

Bridging biological scales by state-space analysis and modeling using molecular, tissue cytometric and physiological data.

Combining data streams across different levels of biological organization such as molecular, cellular, and physiological responses support to a system-wide view in biology. Recently, an unbiased analysis of tissues that provides data-rich descriptors of tissue architecture, cell types, and cell states has become available. As tissues are centrally located in the biological hierarchy, these advancements give rise to a new class of state variables that are critical to elucidate both underlying cellular, molecular and emergent physiological properties. Concepts to statistically identify, correlate, and model relationships across scales are introduced, which rely on a state-space matrix derived by multi-omics data aggregation.

Computational Biology↗
PubMed2006

Exploring molecular networks directly in the cell.

The hierarchy of cell function comprises at least four distinct functional levels: genome, transcriptome, proteome, and toponome. The toponome is the entirety of all protein networks traced out directly as patterns on the single cell level in the natural environment of cells in situ (e.g. tissues). In this work a photonic microscopic robot technology (MELK) capable of tagging and imaging hundreds (and possibly thousands) of different molecular components (e.g. proteins) of morphologically-intact fixed cells and tissue have been developed. MELK data sets represent multidimensional vectors of the topologically determined arrangements of proteins within the cell. The data, assembled in a toponome dictionary of the cell, give rise to a new concept for target and drug lead discovery.

Eukaryotic Cells↗
PubMed2006

Evaluation of a high-content screening fluorescence-based assay analyzing the pharmacological modulation of lipid homeostasis in human macrophages.

BACKGROUND: For understanding cholesterol and phospholipid efflux pathways there is a need for cellular fluorescence-based high-content screens (HCS) to investigated the cholesterol and phospholipid content in human macrophages. METHODS: Making use of fluorescence imaging based on HCS we have developed a tool to evaluate new agents that can act as inducers of cholesterol efflux. The fluorescence assay is based on the different staining patterns of cholesterol-loaded (E-LDL) and deloaded (HDL3) differentiated monocytes by the saturated, fluorescent lipid probe (1,2-dimyristoyl-sn-glycero-3-phospho-ethanolamine)-tetramethyl-rhodamin. RESULTS: Morphologic examination and statistical evaluation of the staining pattern such as gray value, threshold area, shape factor and the spot size distribution provides evidence for a significant pattern change when cholesterol enriched and cholesterol depleted differentiated monocytes were imaged.

Cell Membrane↗
PubMed2006

Imaging spectrometer fundamentals for researchers in the biosciences--a tutorial.

Over the last 2 years there has been a dramatic increase in the number of bioscience laboratories using wavelength dispersive spectroscopy to study in vivo, in situ fluorescence. Transforming spectral information into an image provides a graphic means of mapping localized ionic, molecular, and protein-protein interactions. Spectroscopy also enables fluorophores with overlapping spectral features to be delineation. In this study, we provide the tools that a researcher needs to put into perspective instrumental contributions to a reported spectrum in order to gain greater understanding of the natural emission of the sample. We also show how to deduce the basic capabilities of a spectral confocal system. Finally, we show how to determine the true spectral bandwidth of an object, the illuminated area of a laser-excited object, and what is needed to optimize light throughput.

Animals↗
PubMed2006

Microspectroscopic fluorescence analysis with prism-based imaging spectrometers: review and current studies.

BACKGROUND: Fluorescence imaging spectroscopy is a powerful but under-utilized tool. This article gives perspective on the use of imaging spectroscopy, and provides two examples of imaging spectroscopy done with a prism-based system. The intent is to give insight into the power of imaging spectroscopy when used in combination with other imaging techniques. In particular, studies of intact coral photobleaching and beads designed to show energy transfer are reported. In the bead study, spectroscopic lifetime imaging was performed at each photobleaching step. RESULTS: Spectroscopic photobleaching of the hard coral, Montastrea annularis, revealed two spectral regions. A region in the red portion of the spectrum bleached rapidly while progressively increasing fluorescence was observed over a wide portion of the spectrum. This behavior is consistent with current theories for the role of fluorescent proteins in corals. Following a photobleaching study of beads designed to exhibit energy transfer with imaging spectroscopic fluorescence lifetime imaging microscopy (ISFLIM) allowed unambiguous assignment of fluorescence resonance energy transfer (FRET). The data in this experiment indicated that most of the commonly used markers of FRET would have been inconclusive. The ability of the ISFLIM to look at all regions of the spectrum, particularly the acceptor region, allowed FRET to be assigned. CONCLUSIONS: Fluorescence imaging spectroscopy is a rapidly advancing technology, uniquely suited to the flexible detection of dyes over a wide range of wavelengths.

Animals↗