High-resolution video and digital-enhanced differential interference contrast light microscopy of cell division in budding yeast.
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Cell Cycle↗
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Results for “Microscopy, Interference”
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High-resolution video-enhanced differential interference contrast light microscopy.
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Animals↗
[Proceedings: Interference and polarization microscopy studies on dog hearts following the use of ameroid constrictors].
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Animals↗
Nomarski differential interference contrast--fluorescence microscopy: a new technique designed to improve resolution in fluorescent specimen.
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Animals↗
Microbial growth on agar surfaces studied by incident light differential interference contrast microscopy.
Microbial surface growth on routine opaque agar media was examined by various incident light microscopical techniques. Only differential interference contrast regularly gave good resolution and contrast. The arrangement of units approaching the size of individual bacteria may be judged by low power dry objectives.
Agar↗
Application of Nomarski interference contrast microscopy as a thickness monitor in the preparation of transparent, SiC-based, cross-sectional TEM samples.
Reflected light optical microscopy using a Nomarski prism and a differential interference contrast filter have been employed in concert to achieve a technique that provides an accurate color reference for thickness during the dimpling and ion milling of transparent transmission electron microscopy samples of 6H-SiC(000 1) wafers. The samples had thin films of AIN, GaN, and Au deposited on the SiC substrate. A sequence of variously colored primary and secondary interference bands was observed when the SiC was thinner than 20 microm using an optical microscope. The color bands were correlated with the TEM sample thickness as measured via scanning electron microscopy. The interference contrast was used to provide an indication of the dimpling rate, the ion milling rate, and also the most probable location of perforation, which are useful to reduce sample breakage. The application of pressure during the initial cross-sectional preparation reduced the separation of the two halves of the sample sandwich and resulted in increased shielding of the film surface from ion milling damage.
Journal Article↗
Interference reflection microscopy of adhesion of Amoeba proteus.
A simple method of observing the adhesive behaviour of large cells by means of interference reflection microscope is described, and some observations of monopodial Amoeba proteus are presented. Amoebae may contact with the substratum at any point along the long axis of the cells. Points of contact are usually few, small and temporary. Frequently the cell surface within the contact points oscillates, changing the separation distance from the substratum.
Amoeba↗
Elongation of growth cone filopodia observed with video-enhanced differential interference contrast microscopy.
Dynamics of filopodia configuration from growth cones of neurons isolated from the rat central nervous system were analyzed by using video-enhanced differential interference contrast microscopy. Rapid elongation of filopodia from growth cones and dendrites was observed when high K+ Krebs solution was applied to the growth cones. The maximum speed of the elongation was about 1 micron/s. The filopodia suspended in control Krebs solution moved at random. Retraction of the filopodia was observed within a few minutes after high K+ stimulation.
Animals↗
Reflection interference contrast microscopy combined with scanning force microscopy verifies the nature of protein-ligand interaction force measurements.
The integration of a stand-alone scanning force microscope (SFM) scanner with a reflection interference contrast microscope (RICM) makes it possible to measure directly the separation distance between the SFM probe and the sample surface. The SFM-RICM combination, when applied to the force measurements between ligand-derivatized SFM probe and a protein receptor-derivatized surface, showed that the anomalous force discontinuities often observed for such interacting pairs were indeed a real behavior characteristic of a particular experimental configuration. Apart from small discrepancies due to transient damping, commercially available cantilevers did behave in an ideal mechanical fashion, thus indicating that protein-ligand unbinding events were occurring at distances much larger than their maximum extended length. This external verification of separation distance requires a closer examination of the physical events occurring upon detachment of the surfaces. An alternative interpretation of such force measurements is proposed here in which the protein and/or ligand immobilization chemistry is called into question.
Biophysical Phenomena↗
[Morphologic changes in urine erythrocytes in interference contrast microscopy as an indication of the source of microhematuria].
According to Birch and Fairley (1, 2, 3) it is possible to differentiate glomerular from nonglomerular microhematuria by red cell morphology. Using an interference contrast microscope we examined urine sediments of 85 patients characterized by a microhematuria with more than 8000 red cells/ml urine. Examination of 31 patients with glomerular disease and 27 patients with nonglomerular disease yielded a correlation of 83.5 and 88% respectively between our results and the verified cause of microhematuria. In contrast red cell morphology did not prove any diagnostic favour in 27 patients with renal transplantation.
Diagnosis, Differential↗
The use of urinary red cell morphology to determine the source of hematuria in children.
Urinary red cell morphology has been used to indicate the source of renal tract bleeding. A double blind study was undertaken to evaluate the reliability of urinary red cell morphology in predicting the source of hematuria in a pediatric population. Two independent observers used phase-contrast microscopy, one also using Nomarski differential interference microscopy, to examine 101 urine specimens from 90 pediatric patients with hematuria. 28 cases were subsequently excluded because the clinical diagnosis was uncertain. In the remaining 62 patients the laboratory diagnosis based on the erythrocyte morphology was compared with the primary clinical diagnosis. Observer 1 documented dysmorphic red cells in 40 of 42 patients with glomerulonephritis, and isomorphic red cells in 19 of the 20 cases of non-glomerular bleeding. Observer 2 documented dysmorphic red cells in 39 of the 42 glomerular lesions, and isomorphic red cells in all of the non-glomerular lesions. This represents a sensitivity for predicting glomerular hematuria of 95% for observer 1 and 93% for observer 2, and a specificity of 95% and 100% respectively. There was no definite advantage in the use of Nomarski compared to phase-contrast microscopy. These results confirm the accuracy of this technique in predicting the source of hematuria in pediatric patients.
Child↗
Immunogold and fluorescein immunolabelling of Legionella pneumophila within an aquatic biofilm visualized by using episcopic differential interference contrast microscopy.
Biofilms containing diverse microflora were developed in tap water on glass and polybutylene surfaces. Legionella pneumophila within the biofilms was labelled with monoclonal antibodies and visualized with immunogold or fluorescein isothiocyanate conjugates. Development of a differential interference contrast technique in an episcopic mode enabled simultaneous visualization of the total biofilm flora and gold-labelled legionellae. The legionellae occurred in microcolonies within the biofilm in the absence of amoebae, suggesting that the bacterial consortium was supplying sufficient nutrients to enable legionellae to grow extracellularly within the biofilm.
Antibodies, Monoclonal↗
Application of laser scanning cytometry followed by epifluorescent and differential interference contrast microscopy for the detection and enumeration of Cryptosporidium and Giardia in raw and potable waters.
AIMS: The main goal of this study was to validate a new laser scanning cytometry method (ChemScanRDI) that couples immunofluorescence detection with differential interference contrast (DIC) confirmation, against manual microscopic enumeration of Giardia and Cryptosporidium (oo)cysts. This study also assessed the basic performance of the new Association Française de Normalisation (AFNOR) NF T 90-455 method for Giardia and Cryptosporidium (oo)cyst enumeration with respect to (oo)cyst yield, linearity, repeatability, influence of turbidity and detection limit in raw and potable waters. METHODS AND RESULTS: The new standard method relies on cartridge (Envirocheck) filtration, immunomagnetic separation purification, immunofluorescence staining and detection followed by DIC confirmation. The recovery was 30-50% for both parasites at seeding levels from 30 to 230 (oo)cysts. The method is linear from 0 to around 400 seeded (oo)cysts and the yield does not significantly vary for turbidity levels from 10 to 40 Formazin Nephelometric Units (FNU). The results were obtained using manual microscopic enumeration of the (oo)cysts. The ChemScanRDI yielded counts that were at least equivalent to those obtained using manual microscopy for both parasites in raw and potable water concentrates, for seeding levels of 10-300 or 10-100, respectively. The purification and labelling method proposed by the supplier of theChemScanRDI (Chemunex) reached very similar recoveries to the AFNOR protocol (70-86% in both cases). CONCLUSIONS: Laser scanning cytometry can be used as a more standardized alternative to manual enumeration as part of the new AFNOR standard method. SIGNIFICANCE AND IMPACT OF THE STUDY: By using laser scanning cytometry instead of manual microscopy, laboratories could circumvent the limitations of manual microscopy, namely: low sample throughput, operator subjectivity and operator fatigue. The study further supports the drive to incorporate laser scanning cytometry in the standard methods for Giardia and Cryptosporidium enumeration.
Animals↗
Influence of aqueous fixation on articular surface morphology. A reflected light interference microscope study.
Fresh, unfixed temporomandibular fibrous articular surfaces from baboons were examined by reflected light interference microscopy. The appearances, recorded photographically, were compared with those of joints subjected to aqueous fixation in buffered glutaraldehyde and formalin. Fixation caused readily detectable, measureable changes in articular surface morphology. Reflected light interference microscopy is a valuable technique for the demonstration of the effects of fixation on articular surface morphology.
Aldehydes↗
Differential interference contrast microscopy combined with immunofluorescence: a new method to phenotype eosinophils in situ.
Several surface receptors are expressed on eosinophils in vitro depending on the state of cellular activation, but immunohistochemical studies of eosinophil-related diseases have mostly focused on the number of infiltrating eosinophils as well as extracellular deposits of eosinophil granule proteins. The present investigation showed that eosinophils display a characteristic granular appearance in cryo-sections and cytospins by differential interference contrast (DIC) imaging. This approach appeared to be more reliable for identification of these cells in situ than immunohistochemical labelling of eosinophil granule proteins. Moreover, combined with immunofluorescence microscopy DIC imaging facilitated three-colour immunofluorescence phenotyping of eosinophils.
Cell Count↗
On-stage selection of single round spermatids using a vital, mitochondrion-specific fluorescent probe MitoTracker(TM) and high resolution differential interference contrast microscopy.
The selection of individual round spermatids for round spermatid injection (ROSI), a prerequisite for the successful application of this infertility treatment, has been hampered by the ambiguous definition of a round spermatid and the lack of specific vital and non-vital markers. Using cells from rhesus monkey and bull, we describe a non-invasive method for the on-stage selection of individual round spermatids for ROSI, based on the polarized patterns of mitochondria, visualized in live round spermatid cells by epifluorescence microscopy after incubation with MitoTracker(TM), a vital, mitochondrion-specific fluorescent probe. The correct identification of live round spermatid was confirmed by the presence of the acrosomal granule or acrosomal cap in parallel observations by Nomarski differential interference contrast microscopy. The existence of mitochondrial polarization was first established by the labelling of MitoTracker-tagged round spermatids with spermatid-specific antibodies against proteins of nascent sperm accessory structures combined with antibodies against a nuclear pore complex component, known to disappear at the round spermatid stage. Using an inverted microscope equipped with epifluorescence, the round spermatids can be individually selected from a heterogeneous population of testicular cells labelled with MitoTracker dyes. A major advantage of this approach is that the dyes are incorporated into the paternal mitochondria, destined for rapid elimination after fertilization. In addition, the relatively high excitation and emission wavelengths of MitoTracker dyes are less harmful to DNA after their photon excitation. Before the appropriate clinical testing is conducted, the MitoTracker-based round spermatid selection may be instrumental in the training of clinical staff.
Animals↗
Quantitative phase amplitude microscopy IV: imaging thick specimens.
The ability to image phase distributions with high spatial resolution is a key capability of microscopy systems. Consequently, the development and use of phase microscopy has been an important aspect of microscopy research and development. Most phase microscopy is based on a form of interference. Some phase imaging techniques, such as differential interference microscopy or phase microscopy, have a low coherence requirement, which enables high-resolution imaging but in effect prevents the acquisition of quantitative phase information. These techniques are therefore used mainly for phase visualization. On the other hand, interference microscopy and holography are able to yield quantitative phase measurements but cannot offer the highest resolution. A new approach to phase microscopy, quantitative phase-amplitude microscopy (QPAM) has recently been proposed that relies on observing the manner in which intensity images change with small defocuses and using these intensity changes to recover the phase. The method is easily understood when an object is thin, meaning its thickness is much less than the depth of field of the imaging system. However, in practice, objects will not often be thin, leading to the question of what precisely is being measured when QPAM is applied to a thick object. The optical transfer function formalism previously developed uses three-dimensional (3D) optical transfer functions under the Born approximation. In this paper we use the 3D optical transfer function approach of Streibl not for the analysis of 3D imaging methods, such as tomography, but rather for the problem of analysing 2D phase images of thick objects. We go on to test the theoretical predictions experimentally. The two are found to be in excellent agreement and we show that the 3D imaging properties of QPAM can be reliably predicted using the optical transfer function formalism.
Animals↗
Motility of vinculin-deficient F9 embryonic carcinoma cells analyzed by video, laser confocal, and reflection interference contrast microscopy.
We have studied the motility of wild-type F9 and vinculin-deficient (5.51) mouse embryonal carcinoma cells. F9 cells extended filopodia at a rate of 61 ( +/- 18) nm/s over a distance of 3.18 (+/- 0.29) microns. In contrast, 5.51 cells exhibited filopodia which extended at a similar speed of 57 (+/- 17) nm/s but over a longer distance of 5.10 (+/- 2.14) microns. Cell-substratum contact areas of both cell types were examined by reflection interference contrast microscopy. Wild-type F9 cells had distinct close contacts (dark gray areas) at the cell periphery, whereas 5.51 cells had only a few light gray pinpoint contacts with the substrate. Confocal microscopy showed alpha-actinin to be localized along actin stress fibers in wild-type cells, and in 5.51 cells stress fibers were absent and alpha-actinin was associated with F-actin in the filopodia. beta 1-integrin, talin, and paxillin were concentrated in focal contacts in wild-type cells, but in 5.51 cells beta 1-integrin and talin were in patches under the plasma membrane and paxillin was diffusely distributed in the cytoplasm. We conclude that changes in cell shape and motility of 5.51 compared to wild-type F9 cells are due to the absence of vinculin even though there may be functions of other focal adhesion complex proteins, e.g., talin, linking the actin cytoskeleton to the plasma membrane.
Actinin↗